Synthesis and use for the treatment of osteoporosis

ABSTRACT

Synthetic polypeptide analogs of parathyroid hormone PTH, parathyroid hormone related peptide PTHrp, and of the physiologically active truncated homologs and analogs of PTH and PTHrp, in which amino acid residues (22-31) form an amphipathic α-helix, said residues (22-31) selected from hydrophilic amino acids (Haa) and lipophilic amino acids (Laa) ordered in the sequence: 
     
         Haa(Laa Laa Haa Haa).sub.2 Laa 
    
     and their pharmaceutically acceptable salts are useful for the prophylaxis and treatment of ostoporosis in mammals. Processes for the production of the polypeptides via solid phase and recombinant methods are provided.

This application is a continuation-in-part of U.S. patent applicationSer. No. 07/915,247, filed Jul. 14, 1992 issued as U.S. Pat. No.5,589,452.

BACKGROUND OF THE INVENTION

a) Field of the Invention

This invention relates to novel analogs of parathyroid hormone andparathyroid hormone related peptide, their synthesis by solid phase andrecombinant techniques, and their use for increasing bone mass inmammalian subjects.

b) Description of Related Art

Osteoporosis is the most common form of metabolic bone disease and maybe considered the symptomatic, fracture stage of bone loss (osteopenia).Although osteoporosis may occur secondary to a number of underlyingdiseases, 90% of all cases appear to be idiopathic. Postmenopausal womenare particularly at risk for idiopathic osteoporosis (postmenopausal orType I osteoporosis). Another high risk group for idiopathicosteoporosis is the elderly of either sex (senile or Type IIosteoporosis). Osteoporosis has also been related to corticosteroid use,immobilization or extended bed rest, alcoholism, diabetes, gonadotoxicchemotherapy, hyperprolactinemia, anorexia nervosa, primary andsecondary amenorrhea, and oophorectomy.

In the various forms of osteoporosis, bone fractures, which are theresult of bone loss that has reached the point of mechanical failure,frequently occur. Postmenopausal osteoporosis is characterized byfractures of the wrist and spine, while femoral neck fractures seem tobe the dominant feature of senile osteoporosis.

The mechanism by which bone is lost in osteoporotics is believed toinvolve an imbalance in the process by which the skeleton renews itself.This process has been termed bone remodeling. It occurs in a series ofdiscrete pockets of activity. These pockets appear spontaneously withinthe bone matrix on a given bone surface as a site of bone resorption.Osteoclasts (bone dissolving or resorbing cells) are responsible for theresorption of a portion of bone of generally constant dimension. Thisresorption process is followed by the appearance of osteoblasts (boneforming cells) which then refill with new bone the cavity left by theosteoclasts.

In a healthy adult subject, the rate at which osteoclasts andosteoblasts are formed is such that bone formation and bone resorptionare in balance. However, in osteoporotics an imbalance in the boneremodeling process develops which results in bone being lost at a ratefaster than it is being made. Although this imbalance occurs to someextent in most individuals as they age, it is much more severe andoccurs at a younger age in postmenopausal osteoporotics or followingoophorectomy.

There have been many attempts to treat osteoporosis with the goal ofeither slowing further bone loss or, more desirably, producing a netgain in bone mass. Certain agents, such as estrogen and thebisphosphonates, appear to slow further bone loss in osteoporotics.Agents which slow bone loss, because of the different durations of boneresorption and formation, may appear to increase bone mass (on the orderof 3 to 7%). However, this apparent increase is limited in time, notprogressive, and is due to a decrease in "remodeling space." Inaddition, because of the close coupling between resorption andformation, treatments which impede bone resorption also ultimatelyimpede bone formation.

It has been suggested that treatment with parathyroid hormone (PTH)would lead to both increased bone turnover and a positive calciumbalance. However, human clinical trials have shown that any increase intrabecular bone is offset by a decrease in cortical bone, so that thereis no net increase in total bone.

Hefti, et al. in Clinical Science 62, 389-396 (1982) have reported thatdaily subcutaneous doses of either bPTH(1-84) or hPTH(1-34) increasedwhole body calcium and ash weight of individual bones in both normal andosteoporotic adult female rats.

Liu, et al. in J. Bone Miner. Res. 6:10, 1071-1080 (1991) have notedthat ovariectomy of adult female rats induced a 47% loss in thepercentage of trabecular bone in the proximal tibial metaphysis,accompanied by a significant increase in the number of osteoblasts andtrabecular osteoclasts. Daily subcutaneous injections of hPTH(1-34)completely reversed the loss of trabecular bone and resulted in amountsof trabecular bone exceeding that of sham operated controls. The numberof osteoblasts increased and the number of osteoclasts decreased.

Hock et al. in J. Bone Min. Res. 7:1, 65-71 (1992) have reported thatdaily subcutaneous injections of hPTH(1-34) to healthy adult male ratsfor 12 days increased trabecular and cortical bone calcium and dryweight. Total bone mass, trabecular bone volume, trabecular thicknessand number, and osteoblastic surfaces were increased.

The mammalian parathyroid hormones, e.g. human (hPTH), bovine (bPTH),and porcine (pPTH), are single polypeptide chains of 84 amino acidresidues, with molecular weights of approximately 9500. Biologicalactivity is associated with the N-terminal portion, with residues (1-34)apparently the minimum required.

The N-terminal segment of human PTH differs from the N-terminal segmentof the bovine and porcine hormones by only three and two amino acidresidues, respectively:

    __________________________________________________________________________    hPTH (1-34):    Ser       Val          Ser             Glu                Ile                   Gln                      Leu                         Met                            His                               Asn                                  Leu                                     Gly                                        Lys                                           His                                              Leu    1           5              10             15    Asn       Ser          Met             Glu                Arg                   Val                      Glu                         Trp                            Leu                               Arg                                  Lys                                     Lys                                        Leu                                           Gln                                              Asp                20             25             30    Val       His          Asn             Phe                (SEQ ID NO:1);    bPTH (1-34):    Ala       Val          Ser             Glu                Ile                   Gln                      Phe                         Met                            His                               Asn                                  Leu                                     Gly                                        Lys                                           His                                              Leu    1           5              10             15    Ser       Ser          Met             Glu                Arg                   Val                      Glu                         Trp                            Leu                               Arg                                  Lys                                     Lys                                        Leu                                           Gln                                              Asp                20             25             30    Val       His          Asn             Phe                (SEQ ID NO:2);    pPTH (1-34):    Ser       Val          Ser             Glu                Ile                   Gln                      Leu                         Met                            His                               Asn                                  Leu                                     Gly                                        Lys                                           His                                              Leu    1           5              10             15    Ser       Ser          Leu             Glu                Arg                   Val                      Glu                         Trp                            Leu                               Arg                                  Lys                                     Lys                                        Leu                                           Gln                                              Asp                20             25             30    Val       His          Asn             Phe                (SEQ ID NO:3).    __________________________________________________________________________

The primary function of PTH is to elicit the adaptive changes that serveto maintain a constant concentration of Ca² + in the extracellularfluid. PTH acts on the kidneys to increase tubular reabsorption of Ca² +from the urine, as well as stimulating the conversion of calcifediol tocalcitriol, which is responsible for absorption of Ca² + from theintestines. One prominent effect is to promote the mobilization of Ca² +from bone. PTH acts on bone to increase the rate of resorption of Ca² +and phosphate. PTH stimulates the rate of bone resorption byosteoclasts, increases the rate of differentiation of mesenchymal cellsto osteoclasts, and prolongs the half life of these latter cells. Withprolonged action of PTH the number of bone forming osteoblasts is alsoincreased; thus, the rate of bone turnover and remodeling is enhanced.However, individual osteoblasts appear to be less active than normal.

Rosenblatt, et al. in U.S. Pat. Nos. 4,423,037, 4,968,669 and 5,001,223have disclosed PTH antagonists obtained by the deletion of theN-terminal (1-6) amino acids and the selective replacement of Phe⁷,Met⁸,18, and Gly¹². Tyr³⁴ -NH₂ reportedly increased the activity andstability of these compounds.

Parathyroid hormone-related peptide (PTHrp), a 140+ amino acid protein,and fragments thereof, reproduce the major biological actions of PTH.PTHrp is elaborated by a number of human and animal tumors and othertissues and may play a role in hypercalcemia of malignancy. The sequenceof hPTHrp (1-34) is as follows:

    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Phe                         Phe                            Leu                               His                                  His                                     Leu                                        Ile                                           Ala                                              Glu                20             25             30    Ile       His          Thr             Ala                (SEQ ID NO:4).    __________________________________________________________________________

The sequence homology between hPTH and hPTHrp is largely limited to the13 N-terminal residues, 8 of which are identical; only 1 of 10 aminoacids in the (25-34) receptor binding region of hPTH is conserved inhPTHrp. Conformational similarity may underlie the common activity.Cohen, et al. in J. Biol. Chem. 266:3, 1997-2004 (1991) have suggestedthat much of the sequence of PTH(1-34) and PTHrp(1-34), in particularregions (5-18) and (21-34), assumes an α-helical configuration, whilenoting that there is some question whether this configuration prevailsfor the carboxyl terminal end under physiological conditions. Such asecondary structure may be important for lipid interaction, receptorinteraction, and/or structural stabilization.

We have synthesized analogs of PTH and of PTHrp with the objective ofdeveloping improved therapeutic agents for the restoration of bone massin mammalian subjects, including those afflicted with osteoporosis.

SUMMARY OF THE INVENTION

This invention provides synthetic polypeptide analogs of parathyroidhormone (PTH), parathyroid hormone related peptide (PTHrp), and of thephysiologically active truncated homologs and analogs of PTH and PTHrp,and salts thereof, in which amino acid residues (22-31) form anamphipathic α-helix, said residues (22-31) selected from hydrophilicamino acids (Haa) and lipophilic amino acids (Laa) ordered in thesequence:

    Haa (Laa Laa Haa Haa).sub.2 Laa.

When specific illustrative embodiments of this sequence type areinserted into PTH, PTHrp, and the physiologically active truncatedanalogs and homologs of PTH and PTHrp, the resulting polypeptides areeffective bone remodeling agents.

In one aspect, then, this invention provides analogs of PTH, PTHrp, andthe physiologically active truncated analogs and homologs of PTH andPTHrp, or salts thereof, in which amino acid residues (22-31) form anamphipathic α-helix, the sequence of said residues (22-31) selectedfrom: ##STR1## Xaa¹ and Xaa⁴ are independently Glu, Glu(OCH₃), His, orPhe; Xaa² is Leu or Phe; Xaa⁵ is Lys or His; Xaa⁷ and Xaa¹⁰ areindependently Leu or Ile; Xaa⁸ is Ala, Arg, or Glu; and Xaa⁹ is Lys orGlu (SEQ ID NO:85); preferably ##STR2## Xaa¹ and Xaa⁴ are independentlyGlu, Glu(OCH₃), His, or Phe; Xaa² is Leu or Phe; Xaa⁸ is Glu, Lys, orLys(COCH₂ PEGX) and PEGX is a poly-(ethylene glycol methyl ether)radical of molecular weight 100 to 10,000 (SEQ ID NO:86); preferably,##STR3## Xaa is Glu, Lys, or Lys(COCH₂ PEGX) and PEGX is a poly(ethyleneglycol methyl ether) radical of molecular weight 100 to 10,000 (SEQ IDNO:27); ##STR4##

In another aspect, this invention provides analogs of PTH, PTHrp, and ofthe physiologically active truncated homologs and analogs of PTH andPTHrp, or salts thereof, in which amino acid residues (22-31) form anamphipathic α-helix, the sequence of said residues (22-31) selectedfrom: ##STR5## Xaa is Glu, Lys, or Lys(COCH₂ PEGX) and PEGX is apoly(ethylene glycol methyl ether) radical of molecular weight 100 to10,000 (SEQ ID NO:27); ##STR6##

Also provided are pharmaceutical compositions for the prevention ortreatment of conditions characterized by decreases in bone masscomprising an effective bone mass increasing amount of a polypeptideanalog of parathyroid hormone (PTH), parathyroid hormone related peptide(PTHrp), and of the physiologically active truncated homologs andanalogs of PTH and PTHrp, and salts thereof, in which amino acidresidues (22-31) form an amphipathic α-helix, said residues (22-31)selected from hydrophilic amino acids (Haa) and lipophilic amino acids(Laa) ordered in the sequence:

    Haa (Laa Laa Haa Haa).sub.2 Laa.

in admixture with a pharmaceutically acceptable carrier. Also providedare processes for preparing pharmaceutical compositions which comprisemixing the above described compounds with a pharmaceutically acceptablecarrier.

Further, this invention provides methods for treating mammalianconditions characterized by decreases in bone mass which methodscomprise administering to a mammal in need thereof an effective bonemass increasing amount of a polypeptide analog of PTH, PTHrp, or of aphysiologically active truncated homolog or analog of PTH or PTHrp, or asalt thereof, in which amino acid residues (22-31) form an amphipathicα-helix, said residues (22-31) selected from hydrophilic amino acids(Haa) and lipophilic amino acids (Laa) ordered in the sequence:

    Haa (Laa Laa Haa Haa).sub.2 Laa.

More specifically, this invention provides methods for treatingmammalian conditions characterized by decreases in bone mass whichmethods comprise administering to a mammal in need thereof an effectivebone mass increasing amount of a polypeptide analog of PTH, PTHrp, or ofa physiologically active truncated homolog or analog of PTH or PTHrp, orsalt thereof, in which amino acid residues (22-31) form an amphipathicα-helix, the sequence of said residues (22-31) selected from: (SEQ IDNOS: 85, 86, 26, 27, 28, 29, and 30).

This invention also includes processes for the solid phase synthesis ofpolypeptide analogs of PTH, PTHrp, and of the physiologically activetruncated homologs and analogs of PTH and PTHrp, and salts thereof, inwhich amino acid residues (22-31) form an amphipathic α-helix, saidresidues (22-31) selected from hydrophilic amino acids (Haa) andlipophilic amino acids (Laa) ordered in the sequence:

    Haa (Laa Laa Haa Haa).sub.2 Laa,

which processes comprise sequentially coupling protected amino acids ona suitable resin support, removing the side chain and N.sup.α-protecting groups, and cleaving the polypeptide from the resin.

This invention also encompasses processes for the solid phase synthesisof polypeptide analogs of PTH, pTHrp, and of the physiologically activetruncated homologs and analogs of PTH and PTHrp, and salts thereof, inwhich amino acid residues (22-31) form an amphipathic α-helix, thesequence of said residues (22-31) selected from: (SEQ ID NOS: 85, 86,26, 27, 28, 29, and 30); which processes comprise sequentially couplingprotected amino acids on a suitable resin support, removing the sidechain and N-protecting groups, and cleaving the polypeptide from theresin.

Also included are processes for the recombinant synthesis of polypeptideanalogs of PTH, PTHrp, and of the physiologically active truncatedhomologs and analogs of PTH and PTHrp, and salts thereof, in which aminoacid residues (22-31) form an amphipathic α-helix, said residues (22-31)selected from hydrophilic amino acids (Haa) and lipophilic amino acids(Laa) ordered in the sequence:

    Haa (Laa Laa Haa Haa).sub.2 Laa.

The invention also encompasses DNA sequences, vectors, and plasmids forthe recombinant synthesis of such polypeptide analogs. Specifically, theinvention provides DNA sequences, vectors, and plasmids for therecombinant synthesis of polypeptide analogs of PTH, PTHrp, and of thephysiologically active truncated homologs and analogs of PTH and PTHrp,and salts thereof, in which amino acid residues (22-31) form anamphipathic α-helix, the sequence of said residues (22-31) selectedfrom: (SEQ ID NOS: 85, 86, 26, 27, 28, 29, and 30).

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 presents the DNA sequence and enzyme restriction sites of asynthetic gene coding for a PTHrp(1-34) analog of this invention.

FIG. 2 outlines the preparation of a plasmid incorporating a PTHrp(1-34)analog gene.

FIG. 3 outlines the preparation of a plasmid incorporating two copies ofa PTHrp(1-34) analog gene.

FIG. 4 outlines the preparation of a plasmid incorporating four copiesof a PTHrp(1-34) analog gene.

DETAILED DESCRIPTION OF THE INVENTION Abbreviations and Definitions

The one- and three-letter abbreviations for the various commonnucleotide bases and amino acids are as recommended in Pure Appl. Chem.31, 639-645 (1972) and 40, 277-290 (1974) and the IUPAC-IUB BiochemicalNomenclature Commission and comply with 37 CFR §1.822 (55 FR 18245, May1, 1990). The one- and three-letter abbreviations are as follows:

    ______________________________________    Amino Acid Abbreviations                   Three-letter                             One-letter    Amino Acid     Symbol    Symbol    ______________________________________    Alanine        Ala       A    Arginine       Arg       R    Asparagine     Asn       N    Aspartic Acid  Asp       D    Asn + Asp      Asx    Cysteine       Cys       C    Glutamine      Gln       Q    Glutamic Acid  Glu       E    Gln + Glu      Glx       Z    Glycine        Gly       G    Histidine      His       H    Isoleucine     Ile       I    Leucine        Leu       L    Lysine         Lys       K    Methionine     Met       M    Phenylalanine  Phe       F    Proline        Pro       P    Serine         Ser       S    Threonine      Thr       T    Tryptophan     Trp       W    Tyrosine       Tyr       Y    Valine         Val       V    Other amino acid                   Xaa       X    ______________________________________

The abbreviations represent L-amino acids unless otherwise designated asD- or D,L-. Certain amino acids, both natural and non-natural, areachiral, e.g. glycine. All peptide sequences are presented with theN-terminal amino acid on the left and the C-terminal amino acid on theright.

Further abbreviations for other amino acids and compounds used hereinare:

    ______________________________________    hSer        homoserine    hSerlac     homoserine lactone    Nle         norleucine    PEG2        radical of diethylene glycol methyl                ether, a.k.a. methoxydi(ethyleneoxy),                CH.sub.3 O(CH.sub.2 CH.sub.2 O).sub.2 -, (MW = 119)    PEG5000     radical of poly(ethylene glycol methyl                ether), a.k.a. methoxypoly(ethyleneoxy),                CH.sub.3 O(CH.sub.2 CH.sub.2 O).sub.110 -, (avg. MW = 5000)    PEGX        radical of poly(ethylene glycol methyl                ether), CH.sub.3 O(CH.sub.2 CH.sub.2 O).sub.n -, n = 2-225,                (avg. MW = 100 to 10,000)    ______________________________________

"Hydrophilic amino acid (Haa)" refers to an amino acid having at leastone hydrophilic functional group in addition to those required forpeptide bond formation, such as arginine, asparagine, aspartic acid,glutamic acid, glutamine, histidine, lysine, serine, threonine, andtheir homologs.

"Lipophilic amino acid (Laa)" refers to an uncharged, aliphatic oraromatic amino acid, such as isoleucine, leucine, methionine,phenylalanine, tryptophan, tyrosine, valine, and their homologs.

For the purposes of this invention, alanine is classified as"amphiphilic" i.e., capable of acting as either hydrophilic orlipophilic.

"Physiologically active truncated homolog or analog of PTH or PTHrp"refers to a polypeptide having a sequence comprising less than the fullcomplement of amino acids found in PTH or PTHrp which, however, elicitsa similar physiological response. The truncated PTH or PTHrp need not befully homologous with PTH or PTHrp to elicit a similar physiologicalresponse. PTH(1-34) and PTHrp(1-34) are preferred, but not exclusive,representatives of this group.

"Amphipathic α-helix" refers to the secondary structure exhibited bycertain polypeptides in which the amino acids assume an α-helicalconfiguration having opposing polar and nonpolar faces oriented alongthe long axis of the helix. The possibility of α-helical structure inthe polypeptide of interest may be explored to some extent by theconstruction of a "Schiffer-Edmundson wheel" (M. Schiffer and A. B.Edmundson, Biophys. J. 7, 121 (1967)), of the appropriate pitch andnoting the segregation of the hydrophilic and lipophilic residues onopposite faces of the cylinder circumscribing the helix. Alternatively,empirical evidence, such as circular dichroism or x-ray diffractiondata, may be available indicating the presence of an α-helical region ina given polypeptide. An ideal α-helix has 3.6 amino acid residues perturn with adjacent side chains separated by 100° of arc. Eisenberg etal. in Nature 299:371-374 (1982) and Proc. Nat. Acad. Sci. USA81:140-144 (1984) have combined a hydrophobicity scale with the helicalwheel to quantify the concept of amphipathic helices. The meanhydrophobic moment is defined as the vector sum of the hydrophobicitiesof the component amino acids making up the helix. The followinghydrophobicities for the amino acids are those reported by Eisenberg(1984) as the "consensus" scale:

Ile 0.73; Phe 0.61; Val 0.54; Leu 0.53; Trp 0.37;

Met 0.26 Ala 0.25; Gly 0.16; Cys 0.04; Tyr 0.02;

Pro -0.07; Thr -0.18; Ser -0.26; His -0.40; Glu -0.62;

Asn -0.64; Gln -0.69; Asp -0.72; Lys -1.10; Arg -1.76.

The hydrophobic moment, μ_(H), for an ideal α-helix having 3.6 residuesper turn (or a 100° arc (=360°/3.6) between side chains), may becalculated from:

    μ.sub.H = (ε H.sub.N sin δ(N-1)).sup.2 +(ε H.sub.N cos δ(N-1).sup.2 !.sup.1/2,

where H_(N) is the hydrophobicity value of the N^(th) amino acid and thesums are taken over the N amino acids in the sequence with periodicityδ=100°. The hydrophobic moment may be expressed as the mean hydrophobicmoment per residue by dividing μ_(H) by N to obtain <μ_(H) >. A value of<μ_(H) > at 100°±20° of about 0.20 or greater is suggestive ofamphipathic helix formation. The <A_(H) > values at 100° for hPTHrp(22-31) and hPTH (22-31) are 0.19 and 0.37, respectively.

Cornett, et al., in J. Mol. Biol., 195:659-685 (1987) have furtherextended the study of amphiphathic α-helices by introducing the"amphipathic index" as a predictor of amphipathicity. They concludedthat approximately half of all known α-helices are amphipathic, and thatthe dominant frequency is 97.5° rather than 100°, with the number ofresidues per turn being closer to 3.7 than 3.6. While such refinementsare scientifically interesting, the basic approach of Eisenberg, et al.is sufficient to classify a given sequence as amphipathic, particularlywhen one is designing a sequence ab initio to form an amphipathicα-helix.

A substitute amphipathic α-helical amino acid sequence may lack homologywith the sequence of a given segment of a naturally occurringpolypeptide but elicits a similar secondary structure, i.e. an α-helixhaving opposing polar and nonpolar faces, in the physiologicalenvironment. Replacement of the naturally occurring amino acid sequencewith an alternative sequence may beneficially affect the physiologicalactivity, stability, or other properties of the altered parentpolypeptide. Guidance as to the design and selection of such sequencesis provided in J. L. Krstenansky, et al., FEBS Letters 242:2, 409-413(1989), and J. P. Segrest, et al. Proteins: Structure, Function, andGenetics 8:103-117 (1990) among others.

The ten amino acid amphipathic α-helix of this invention has theformula:

    Haa (Laa Laa Haa Haa).sub.2 Laa

wherein the Haa's are selected from the group of hydrophilic amino acidsand the Laa's are selected from the group of lipophilic amino acids, asdefined above. Assuming an idealized α-helix, residues 1, 4, 5, 8, and 9are distributed along one face (A) of the helix within about a 140° arcof each other, while residues 2, 3, 6, 7, and 10 occupy an opposing 140°arc on the other face (B) of the helix. Preferably, all the residues onone face are of the same polarity while all those on the other face areof the opposite polarity, i.e., if face A is all hydrophilic, face B isall lipophilic and vice versa. The skilled artisan will recognize thatwhile the helices of this invention are described by

    Haa(Laa Laa Haa Haa).sub.2 Laa,

the reverse sequence,

    Laa (Haa Haa Laa Laa).sub.2 Haa

will also meet the residue distribution criteria and is an equivalentdescriptor of the helices of this invention.

Alanine may be substituted for either hydrophilic or lipophilic aminoacids, since Ala can reside readily on either face of an amphipathicα-helix, although Ala₁₀ does not form an amphipathic α-helix. Generally,proline, cysteine, and tyrosine are not used; however, their presenceand other random errors in the sequence may be tolerated, e.g. ahydrophilic residue on the lipophilic face, as long as the remainingamino acids in the segment substantially conform to the hydrophilicface--lipophilic face division. A convenient method for determining if asequence is sufficiently amphipathic to be a sequence of this inventionis to calculate the mean hydrophobic moment, as defined above. If thepeak mean moment per residue at 100°±20° exceeds about 0.20, then thesequence will form an amphipathic helix and is a sequence of thisinvention.

For example, the mean hydrophobic moment per residue at 100° for (SEQ IDNO: 26), Xaa=Glu, is calculated as follows:

    ______________________________________    A.A.  H.sub.N                 δ (N - 1)                            H sin δ (N - 1)                                     H cos δ (N - 1)    ______________________________________    E     -.62    0         0        -.62    L     .53    100        .52      -.17    L     .53    200        -.18     -.50    E     -.62   300        .34      -.31    K     -1.1   400        -.70     -.85    L     .53    500        .34      -.41    L     .53    600        -.46     -.27    E     -.62   700        .21      -.58    K     -1.1   800        -1.08    -.19    L     .53    900        0        -.53                            Σ = 0.81                                     Σ = -4.43    ______________________________________     μ.sub.H =  (0.81).sup.2 + (-4.43).sup.2 !.sup.1/2 = 4.50 < μH > =     4.50/10 = 0.45

For this sequence, the mean peak hydrophobic moment occurs at 92° andhas a value of 0.48.

In applying this concept to parathyroid hormone and parathyroid hormonerelated peptide, it was hypothesized that either or both regions (7-16)and (22-31) may exhibit α-helical secondary structure and could bereplaced with a non-homologous sequence having similar structuraltendencies, without loss of biological activity or induction ofimmunoreaction.

PREFERRED EMBODIMENTS

In one aspect, this invention provides analogs of PTH, PTHrp, and thephysiologically active truncated analogs and homologs of PTH and PTHrp,or salts thereof, in which amino acid residues (22-31) form anamphipathic α-helix, the sequence of said residues (22-31) selectedfrom: ##STR7## Xaa¹ and Xaa⁴ are independently Glu, Glu(OCH₃), His, orPhe; Xaa² is Leu or Phe; Xaa⁵ is Lys or His; Xaa⁷ and Xaa¹⁰ areindependently Leu or Ile; Xaa⁸ is Ala, Arg, or Glu; and Xaa⁹ is Lys orGlu (SEQ ID NO: 85); preferably ##STR8## Xaa¹ and Xaa⁴ are independentlyGlu, Glu(OCH₃), His, or Phe; Xaa² is Leu or Phe; Xaa⁸ is Glu, Lys, orLys(COCH₂ PEGX) and PEGX is a poly-(ethylene glycol methyl ether)radical of molecular weight 100 to 10,000 (SEQ ID NO:86); preferably,##STR9## Xaa is Glu, Lys, or Lys(COCH₂ PEGX) and PEGX is a poly(ethyleneglycol methyl ether) radical of molecular weight 100 to 10,000 (SEQ IDNO:27); ##STR10##

In another aspect, this invention provides analogs of PTH, PTHrp, andthe physiologically active truncated analogs and homologs of PTH andPTHrp, or salts thereof, of the formula:

    Xaa.sup.1 Xaa.sup.2 Xaa.sup.3 Xaa.sup.4 Xaa.sup.5 Xaa.sup.6 Xaa.sup.7 Leu His Xaa.sup.10 Xaa.sup.11 Gly Xaa.sup.13 Ser Ile Gln Xaa.sup.17 Leu Xaa.sup.19 Xaa.sup.20 Xaa.sup.21 Xaa.sup.22-31 Xaa.sup.32 Xaa.sup.33 Xaa.sup.34 Xaa.sup.35 Xaa.sup.36 Xaa.sup.37 Xaa.sup.38 Term,

wherein:

Xaa¹ is absent or is Ala;

Xaa² is absent or is Val;

Xaa³ is absent or is Ser;

Xaa⁴ is absent or is Glu or Glu(OCH₃);

Xaa⁵ is absent or is His or Ala;

Xaa⁶ is absent or is Gln;

Xaa⁷ is absent or is Leu;

Xaa¹⁰ and Xaa¹⁷ are independently Asp or Asp(OCH₃);

Xaa¹ l is Lys, Arg, or Leu;

Xaa¹³ is Lys, Arg, Tyr, Cys, Leu, Cys(CH₂ CONH(CH₂)₂ NH(biotinyl)),Lys(7-dimethylamino-2-oxo-2H-1-benxopyran-4-acetyl), orLys(dihydrocinnamoyl);

Xaa²⁰ is Arg or Leu;

Xaa¹⁹ and Xaa²¹ are independently Lys, Ala, or Arg;

Xaa²²⁻³¹ is selected from (SEQ ID NOS:26, 27, 28, 29, or 30);

Xaa³² is His, Pro, or Lys;

Xaa³³ is absent, or is Pro, Thr, Glu, or Ala;

Xaa³⁴ is absent, or is Pro, Arg, Met, Ala, hSer, hSer lactone, Tyr, Leu,or 1,4-diaminobutyryl lactam;

Xaa³⁵ is absent or is Pro, Glu, Ser, Ala, or Gly;

Xaa³⁶ is absent or is Ala, Arg, or Ile;

Xaa³⁷ is absent or is Arg, Trp, or 3-(-2-naphthyl)-L-alanine;

Xaa³⁸ is absent or is Ala or hSer or Xaa³⁸⁻⁴² is Thr Arg Ser Ala Trp;

and Term is OR or NR₂ where each R is independently H, (C₁ -C₄)alkyl orphenyl(C₁ -C₄)alkyl; and the pharmaceutically acceptable salts thereof.

In yet another aspect this invention includes polypeptide analogs of thephysiologically active truncated homolog hPTHrp(1-34), as shown inFormula (I):

    Ala Val Ser Glu Xaa.sup.5 Gln Leu Leu His Asp Xaa.sup.1 l Gly Xaa.sup.13 Ser Ile Gln Asp Leu Xaa.sup.19 Arg Xaa.sup.21 Xaa.sup.22-31 Xaa.sup.32 Xaa.sup.33 Xaa.sup.34 Term,

wherein:

Xaa⁵ is His or Ala;

Xaa¹¹ and Xaa¹³ are independently Lys, Arg, or Leu;

Xaa¹⁹ and Xaa²¹ are independently Ala or Arg;

Xaa²²⁻³¹ is selected from: ##STR11## Xaa is Glu, Lys, or Lys(COCH₂ PEGX)and PEGX is a poly(ethylene glycol methyl ether) radical of molecularweight 100 to 10,000 (SEQ ID NO:27); ##STR12## Xaa³² is His or Lys;Xaa³³ is Thr, Glu, or Ala;

Xaa³⁴ is Ala, hSer, Tyr, or Leu;

and Term is Gly Arg Arg, lactone, OH or NR₂, where each R is H or (C₁-C₄) alkyl; and their pharmaceutically acceptable salts. (Formula I)

A more specific aspect of the invention includes those polypeptides ofFormula (I) wherein Xaa²²⁻³¹ is (SEQ ID NO:26), for which <A_(H) > at100° exceeds 0.45. A still more specific aspect of the inventionincludes those Formula (I) polypeptides wherein Xaa²²⁻³¹ is (SEQ IDNO:26); Xaa¹ l and Xaa¹³ are both Lys; and Xaa¹⁹ and Xaa²¹ are both Arg.

Representative polypeptides include, but are not limited to:

    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Arg                                              Lys                20             25             30    Leu       His          Thr             Ala                OH (SEQ ID NO:5);    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                OH (SEQ ID NO:6);    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2                   (SEQ ID NO:7);    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             hSer                NH.sub.2                   (SEQ ID NO:8);    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             hSerlac  (SEQ ID NO:9);    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                Gly                   Arg                      Arg                         OH (SEQ ID NO:10); and                35    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       Lys          Glu             Leu                NH.sub.2                   (SEQ ID NO:11).    __________________________________________________________________________

Another aspect of this invention includes those polypeptides of Formula(I) wherein Xaa²²⁻³¹ is (SEQ ID NO:26) ; Xaa¹¹ and Xaa¹³ are both Lys;and one of Xaa¹⁹ and Xaa²¹ is Arg and the other is Ala. Representativepolypeptides of this subgenus include, but are not limited to:

    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Ala                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2                   (SEQ ID NO:12) and    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Ala                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2                   (SEQ ID NO:13).    __________________________________________________________________________

In another aspect this invention includes those polypeptides of Formula(I) wherein Xaa²²⁻³¹ is (SEQ ID NO:26); one of Xaa¹¹ and Xaa¹³ is Leuand the other is Lys; and Xaa¹⁹ and Xaa²¹ are both Arg. Representativepolypeptides of this subgenus include, but are not limited to:

    __________________________________________________________________________    Ala       Val          Ser             Glu                Ala                   Gln                      Leu                         Leu                            His                               Asp                                  Leu                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Ala             Leu                OH (SEQ ID NO:14).    __________________________________________________________________________

In another aspect this invention includes those polypeptides of Formula(I) wherein Xaa²²⁻³¹ is (SEQ ID NO:27), for which <μ_(H) > at 100°exceeds 0.50. A further aspect of this invention includes those Formula(I) polypeptides wherein Xaa²²⁻³¹ is (SEQ ID NO:27); Xaa¹¹ and Xaa¹³ areboth Lys or both Arg; and Xaa¹⁹ and Xaa²¹ are both Arg. Representativepolypeptides of this subgenus include, but are not limited to:

    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Glu                                              Arg                20             25             30    Leu       His          Thr             Ala                OH (SEQ ID NO:i5);    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Arg                                     Gly                                        Arg                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Glu                                              Arg                20             25             30    Leu       His          Thr             Ala                OH (SEQ ID NO:16);    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Arg                                     Gly                                        Arg                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Lys                                              Arg                20             25             30    Leu       His          Thr             Ala                OH (SEQ ID NO:17);    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Arg                                     Gly                                        Arg                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                            20          25    Lys(COCH.sub.2 PEG2)             Arg                Leu                   His                      Thr                         Ala                            OH (SEQ ID NO:18); and                         30    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Arg                                     Gly                                        Arg                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                20             25    Lys(COCH.sub.2 PEG5000)                Arg                   Leu                      His                         Thr                            Ala                               OH (SEQ ID NO:19).                30    __________________________________________________________________________

In another aspect this invention includes polypeptides of Formula (I)wherein Xaa²²⁻³¹ (SEQ ID NO:28), for which <μ_(H) > at 100° is about0.25. Representative polypeptides of this subgenus include, but are notlimited to:

    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Ala                         Leu                            Ala                               Glu                                  Ala                                     Leu                                        Ala                                           Glu                                              Ala                20             25             30    Leu       His          Thr             Ala                NH.sub.2                   (SEQ ID NO:20).    __________________________________________________________________________

In another aspect this invention includes polypeptides of Formula (I)wherein Xaa²²⁻³¹ is (SEQ ID NO:29), for which <μ_(H) > at 100° is about0.28. Representative polypeptides of this subgenus include, but are notlimited to:

    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Ser                         Leu                            Leu                               Ser                                  Ser                                     Leu                                        Leu                                           Ser                                              Ser                20             25             30    Leu       His          Thr             Ala                NH.sub.2                   (SEQ ID NO:21).    __________________________________________________________________________

In another aspect this invention includes polypeptides of Formula (I)wherein Xaa²²⁻³¹ is (SEQ ID NO:30), for which <μ_(H) > at 100° is about0.29. Representative polypeptides of this subgenus include, but are notlimited to:

    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Ala                         Phe                            Tyr                               Asp                                  Lys                                     Val                                        Ala                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2                   (SEQ ID NO:22).    __________________________________________________________________________

Still another aspect of this invention includes polypeptide analogs ofthe physiologically active truncated homolog bPTH(1-34), as shown inFormula (II):

    Xaa.sup.1 Val Ser Glu Ile Gln Xaa.sup.7 Xaa.sup.8 His Asn Leu Gly Lys His Leu Xaa.sup.16 Ser Xaa.sup.18 Xaa.sup.19 Arg Xaa.sup.2 l Xaa.sup.22-31 His Asn Xaa.sup.34 Term,

wherein:

Xaa¹ is Ser or Ala;

Xaa⁷ is Leu or Phe;

Xaa⁸ is Met or Nle;

Xaa¹⁶ is Asn or Ser;

Xaa¹⁸ is Leu, Met, or Nle;

Xaa¹⁹ is Glu or Arg;

Xaa²¹ is Val or Arg;

Xaa²²⁻³¹ is selected from (SEQ ID NO:26, 27, 28, 29, and 30);

Xaa³⁴ is Phe or Tyr;

Term is OH or NR₂, where each R is H or (C₁ -C₄)alkyl; and thepharmaceutically acceptable salts thereof. (Formula II)

Representative polypeptides include, but are not limited to:

    __________________________________________________________________________    Ala       Val          Ser             Glu                Ile                   Gln                      Phe                         Nle                            His                               Asn                                  Leu                                     Gly                                        Lys                                           His                                              Leu    1           5              10             15    Ser       Ser          Nle             Glu                Arg                   Val                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Asn             Tyr                NH.sub.2                   (SEQ ID NO:23) and    Ala       Val          Ser             Glu                Ile                   Gln                      Phe                         Nle                            His                               Asn                                  Leu                                     Gly                                        Lys                                           His                                              Leu    1           5              10             15    Ser       Ser          Nle             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Asn             Tyr                NH.sub.2                   (SEQ ID NO:24).    __________________________________________________________________________

In still another aspect of this invention, it has surprisingly beenfound that homologs and analogs of PTH and PTHrP having less than 34amino acids are also potent bone remodeling agents. These compounds areof general formula:

    Ala Val Ser Glu Xaa.sup.5 Gln Leu Leu His Asp Xaa.sup.11 Gly Xaa.sup.13 Ser Ile Gln Asp Leu Xaa.sup.19 Arg Xaa.sup.21 Xaa.sup.22-31 Xaa.sup.32 Xaa.sup.33 Xaa.sup.34 Term,

Representative polypeptides include, but are not limited to:

Compound 41: AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHP-NH₂ (SEQ ID NO:55)

    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Pro             NH.sub.2                (SEQ ID NO:55).    __________________________________________________________________________

Physical Data

    ______________________________________    m.p. 142.8-166.1° C.    α!.sub.D.sup.25   -53.80 (c 0.38,    H.sub.2 O)    FAB (C.sub.173 H.sub.295 N.sub.55 O.sub.49):  M+H!.sup.+  3929    AAA:    Asx    2.0 (2)  Glx  5.7 (6)                                        Ser  1.8 (2)            His    3.0 (3)  Gly  1.1 (1)                                        Ala  0.9 (1)            Arg    2.8 (3)  Val  1.2 (1)                                        Ile  0.9 (1)            Leu    7.4 (8)  Lys  4.4 (4)                                        Pro  0.9 (1)    ______________________________________

Compound 42: AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LP-NH₂ (SEQ ID NO:56)

    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       Pro          NH.sub.2             (SEQ ID NO:56).    __________________________________________________________________________

Physical Data

    ______________________________________    m.p. 161.0-177.0° C.    α!.sub.D.sup.25   -61.97 (c 0.19,    H.sub.2 O)    FAB (C.sub.167 H.sub.288 N.sub.52 O.sub.48):  M+H!.sup.+  3792.0    AAA:    Asx    2.2 (2)  Glx  5.9 (6)                                        Ser  1.9 (2)            His    2.1 (2)  Gly  1.1 (1)                                        Ala  1.0 (1)            Arg    3.0 (3)  Val  1.1 (1)                                        Ile  1.0 (1)            Leu    7.9 (8)  Lys  4.3 (4)                                        Pro  0.9 (1)    ______________________________________

The skilled artisan will appreciate that numerous permutations of thepolypeptide analogs may be synthesized which will possess the desirableattributes of those described herein provided that an amino acidsequence having a mean hydrophobic moment per residue at 100°±20°greater than about 0.20 is inserted at positions (22-31).

Classical Synthesis of the Polypeptides

The polypeptides of the instant invention may be synthesized by methodssuch as those set forth in J. M. Stewart and J. D. Young, Solid PhasePeptide Synthesis, 2nd ed., Pierce Chemical Co., Rockford, Ill. (1984)and J. Meienhofer, Hormonal Proteins and Peptides, Vol. 2, AcademicPress, New York, (1973) for solid phase synthesis and E. Schroder and K.Lubke, The Peptides, Vol. 1, Academic Press, New York, (1965) forsolution synthesis.

In general, these methods involve the sequential addition of protectedamino acids to a growing peptide chain. Normally, either the amino orcarboxyl group of the first amino acid and any reactive side chain groupare protected. This protected amino acid is then either attached to aninert solid support, or utilized in solution, and the next amino acid inthe sequence, also suitably protected, is added under conditionsamenable to formation of the amide linkage. After all the desired aminoacids have been linked in the proper sequence, protecting groups and anysolid support are removed to afford the crude polypeptide. Thepolypeptide is desalted and purified, preferably chromatographically, toyield the final product.

A preferred method of preparing the analogs of the physiologicallyactive truncated polypeptides, having fewer than about forty aminoacids, involves solid phase peptide synthesis. In this method theα-amino (N.sup.α) functions and any reactive side chains are protectedby acid- or base-sensitive groups. The protecting group should be stableto the conditions of peptide linkage formation, while being readilyremovable without affecting the extant polypeptide chain. Suitablea-amino protecting groups include, but are not limited tot-butoxycarbonyl (Boc), benzyloxycarbonyl (Cbz),o-chlorobenzyloxycarbonyl, biphenylisopropyloxycarbonyl,t-amyloxycarbonyl (Amoc), isobornyloxycarbonyl,α,α-dimethyl-3,5-dimethoxybenzyloxy-carbonyl, o-nitrophenylsulfenyl,2-cyano-t-butoxycarbonyl, 9-fluorenylmethoxycarbonyl (Fmoc) and thelike, preferably t-butoxycarbonyl (Boc). Suitable side chain protectinggroups include, but are not limited to: acetyl, benzyl (Bzl),benzyloxymethyl (Bom), o-bromobenzyloxycarbonyl, t-butyl,t-butyldimethylsilyl, 2-chlorobenzyl (Cl-z), 2,6-dichlorobenzyl,cyclohexyl, cyclopentyl, isopropyl, pivalyl, tetrahydropyran-2-yl, tosyl(Tos), trimethylsilyl, and trityl.

In solid phase synthesis, the C-terminal amino acid is first attached toa suitable resin support. Suitable resin supports are those materialswhich are inert to the reagents and reaction conditions of the stepwisecondensation and deprotection reactions, as well as being insoluble inthe media used. Examples of commercially available resins includestyrene/divinylbenzene resins modified with a reactive group, e.g.,chloromethylated co-poly-(styrene-divinylbenzene), hydroxymethylatedco-poly-(styrene-divinylbenzene), and the like. Benzylated,hydroxy-methylated phenylacetamidomethyl (PAM) resin is preferred. Whenthe C-terminus of the compound is an amide, a preferred resin isp-methylbenzhydrylamino-co-poly(styrene-divinyl-benzene) resin.

Attachment to the PAM resin may be accomplished by reaction of theN.sup.α -protected amino acid, preferably the Boc-amino acid, as itsammonium, cesium, triethylammonium, 1,5-diazabicyclo- 5.4.0!undec-5-ene,tetramethylammonium, or similar salt in ethanol, acetonitrile,N,N-dimethylformamide (DMF), and the like, preferably the cesium salt inDMF, with the resin at an elevated temperature, for example betweenabout 40° and 60° C., preferably about 50° C., for from about 12 to 72hours, preferably about 48 hours.

The N.sup.α -Boc-amino acid may be attached to the benzhydrylamine resinby means of, for example, an N,N'-diisopropylcarbodiimide(DIC)/1-hydroxybenzotriazole (HOBt) mediated coupling for from about 2to about 24 hours, preferably about 2 hours at a temperature of betweenabout 10° and 50° C., preferably 25° C. in a solvent such asdichloromethane or dimethylformamide, preferably dichloromethane.

The successive coupling of protected amino acids may be carried out bymethods well known in the art, typically in an automated peptidesynthesizer. Following neutralization with triethylamine or similarbase, each protected amino acid is preferably introduced inapproximately 1.5 to 2.5 fold molar excess and the coupling carried outin an inert, nonaqueous, polar solvent such as dichloromethane, DMF, ormixtures thereof, preferably in dichloromethane at ambient temperature.Representative coupling agents are N,N'-dicyclohexylcarbodiimide (DCC),N,N'-diisopropylcarbodiimide (DIC) or other carbodiimide, either aloneor in the presence of 1-hydroxybenzotriazole (HOBt), O-acyl ureas,benzotriazol-1-yl-oxytris(pyrrolidino)phosphonium hexafluorophosphate(PyBop), N-hydroxysuccinimide, other N-hydroxyimides, or oximes.Alternatively, protected amino acid active esters (e.g. p-nitrophenyl,pentafluorophenyl and the like) or symmetrical anhydrides may be used.

At the end of the solid phase synthesis the fully protected peptide isremoved from the resin. When the linkage to the resin support is of thebenzyl ester type, cleavage may be effected by means of aminolysis withan alkylamine or fluoroalkylamine for peptides with an alkylamideC-terminus, or by aminolysis with, for example, ammonia/methanol orammonia/ethanol for peptides with an unsubstituted amide C-terminus, ata temperature between about -10° and 50° C., preferably about 25° C.,for between about 12 and 24 hours, preferably about 18 hours. Peptideswith a hydroxy C-terminus may be cleaved by HF or other strongly acidicdeprotection regimen or by saponification. Alternatively, the peptidemay be removed from the resin by transesterification, e.g., withmethanol, followed by aminolysis or saponification. The protectedpeptide may be purified by silica gel chromatography.

The side chain protecting groups may be removed from the peptide bytreating the aminolysis product with, for example, anhydrous liquidhydrogen fluoride in the presence of anisole or other carbonium ionscavenger, treatment with hydrogen fluoride/pyridine complex, treatmentwith tris(trifluoroacetyl)boron and trifluoroacetic acid, by reductionwith hydrogen and palladium on carbon or polyvinylpyrrolidone, or byreduction with sodium in liquid ammonia, preferably with liquid hydrogenfluoride and anisole at a temperature between about -10° and +10° C.,preferably at about 0° C., for between about 15 minutes and 2 hours,preferably about 1.5 hours. For peptides on the benzhydrylamine resin,the resin cleavage and deprotection steps may be combined in a singlestep utilizing liquid hydrogen fluoride and anisole as described above.

The solution may be desalted (e.g. with BioRad AG-3® anion exchangeresin) and the peptide purified by a sequence of chromatographic stepsemploying any or all of the following types: ion exchange on a weaklybasic resin in the acetate form; hydrophobic adsorption chromatographyon underivatized co-poly(styrene-divinylbenzene), e.g. Amberlite® XAD;silica gel adsorption chromatography; ion exchange chromatography oncarboxymethylcellulose; partition chromatography, e.g. on Sephadex®G-25; countercurrent distribution; or high performance liquidchromatography (HPLC), especially reversed-phase HPLC on octyl- oroctadecylsilylsilica (ODS) bonded phase column packing.

Thus, another aspect of the present invention relates to processes forpreparing polypeptides and pharmaceutically acceptable salts thereofwhich processes comprise sequentially condensing protected amino acidson a suitable resin support, removing the protecting groups and resinsupport, and purifying the product, to afford analogs of thephysiologically active truncated homologs and analogs of PTH and PTHrp,preferably of PTH(1-34) and PTHrp(1-34), in which the amino acids atpositions (22-31) form an amphipathic α-helical peptide sequence, asdefined above.

Recombinant Synthesis of the Polypeptides

Alternatively, the polypeptides of this invention may be prepared bycloning and expression of a gene encoding for the desired polypeptide.In this process, a plasmid containing the desired DNA sequence isprepared and inserted into an appropriate host microorganism, typicallya bacteria, such as E. coli, or a yeast, such as Saccharomycescerevisiae, inducing the host microorganism to produce multiple copiesof the plasmid, and so of the cDNA encoding for the polypeptide analogsof this invention.

First, a synthetic gene coding for the selected PTH or PTHrp analog isdesigned with convenient restriction enzyme cleavage sites to facilitatesubsequent alterations. Polymerase chain reaction (PCR), as taught byMullis in U.S. Pat. Nos. 4,683,195 and 4,683,202, may be used to amplifythe sequence.

The amplified synthetic gene may be isolated and ligated to a suitableplasmid, such as a Trp LE plasmid, into which four copies of the genemay be inserted in tandem. Preparation of Trp LE plasmids is describedin U.S. Pat. No. 4,738,921 and European Patent Publication No. 0212532.Trp LE plasmids generally produce 8-10 times more protein than Trp Eplasmids. The multi-copy gene may then be expressed in an appropriatehost, such as E. coli or S. cerevisiae.

The specific expression vector used herein was Trp LE 18 Prot (Ile³,Pro⁵) containing the following elements: a pBR322 fragment (EcoRI-BamHI)containing the ampicillin resistant gene and the plasmid origin ofreplication; an EcoRI-SacII fragment containing the trp promoter and thetrpE gene; an HIV protease (Ile³,Pro⁵) gene fragment (SacII-HindIII); abGRF gene fragment (HindIII-BamHI); and a transcription terminator fromE. coli rpoc gene. The HIV protease and bGRF gene fragments are notcritical and may be replaced with other coding sequences, if desired.

The expressed multimeric fusion proteins accumulate intracellularly intostable inclusion bodies and may be separated by centrifugation from therest of the cellular protein. The isolated fusion protein is convertedto the monomeric PTH or PTHrp analog and may be purified by cationexchange and/or reverse phase HPLC.

Alternative methods of cloning, amplification, expression, andpurification will be apparent to the skilled artisan. Representativemethods are disclosed in Maniatis, et al., Molecular Cloning, aLaboratory Manual, 2nd Ed., Cold Spring Harbor Laboratory (1989).

Utility and Administration

The polypeptides of this invention are useful for the prevention andtreatment of a variety of mammalian conditions manifested by loss ofbone mass. In particular, the compounds of this invention are indicatedfor the prophylaxis and therapeutic treatment of osteoporosis andosteopenia in humans.

In general, the polypeptides of this invention, or salts thereof, areadministered in amounts between about 0.002 and 1 μg/kg body weight perday, preferably from about 0.04 to about 0.2 μg/kg body weight per day.For a 50 kg human female subject, the daily dose of active ingredient isfrom about 0.1 to about 50 μgs, preferably from about 2.0 to about 10μgs. In other mammals, such as horses, dogs, and cattle, higher dosesmay be required. This dosage may be delivered in a conventionalpharmaceutical composition by a single administration, by multipleapplications, or via controlled release, as needed to achieve the mosteffective results, preferably one or more times daily by injection.

The selection of the exact dose and composition and the most appropriatedelivery regimen will be influenced by, inter alia, the pharmacologicalproperties of the selected polypeptide, the nature and severity of thecondition being treated, and the physical condition and mental acuity ofthe recipient.

Representative delivery regimens include oral, parenteral (includingsubcutaneous, intramuscular and intravenous), rectal, buccal (includingsublingual), pulmonary, transdermal, and intranasal.

Pharmaceutically acceptable salts retain the desired biological activityof the parent polypeptide without toxic side effects. Examples of suchsalts are (a) acid addition salts formed with inorganic acids, forexample hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoricacid, nitric acid and the like; and salts formed with organic acids suchas, for example, acetic acid, oxalic acid, tartaric acid, succinic acid,maleic acid, fumaric acid, gluconic acid, citric acid, malic acid,ascorbic acid, benzoic acid, tannic acid, pamoic acid, alginic acid,polyglutamic acid, naphthalenesulfonic acids, naphthalene disulfonicacids, polygalacturonic acid and the like; (b) base addition saltsformed with polyvalent metal cations such as zinc, calcium, bismuth,barium, magnesium, aluminum, copper, cobalt, nickel, cadmium, and thelike; or with an organic cation formed from N,N'-dibenzylethylenediamineor ethylenediamine; or (c) combinations of (a) and (b), e.g., a zinctannate salt and the like.

A further aspect of the present invention relates to pharmaceuticalcompositions comprising as an active ingredient a polypeptide of thepresent invention, or pharmaceutically acceptable salt thereof, inadmixture with a pharmaceutically acceptable, non-toxic carrier. Asmentioned above, such compositions may be prepared for parenteral(subcutaneous, intramuscular or intravenous) administration,particularly in the form of liquid solutions or suspensions; for oral orbuccal administration, particularly in the form of tablets or capsules;for pulmonary or intranasal administration, particularly in the form ofpowders, nasal drops or aerosols; and for rectal or transdermaladministration.

The compositions may conveniently be administered in unit dosage formand may be prepared by any of the methods well-known in thepharmaceutical art, for example as described in Remington'sPharmaceutical Sciences, 17th ed., Mack Publishing Company, Easton, Pa.,(1985). Formulations for parenteral administration may contain asexcipients sterile water or saline, alkylene glycols such as propyleneglycol, polyalkylene glycols such as polyethylene glycol, oils ofvegetable origin, hydrogenated naphthalenes and the like. For oraladministration, the formulation can be enhanced by the addition of bilesalts or acylcarnitines. Formulations for nasal administration may besolid and may contain excipients, for example, lactose or dextran, ormay be aqueous or oily solutions for use in the form of nasal drops ormetered spray. For buccal administration typical excipients includesugars, calcium stearate, magnesium stearate, pregelatinated starch, andthe like.

When formulated for nasal administration, the absorption across thenasal mucous membrane may be enhanced by surfactant acids, such as forexample, glycocholic acid, cholic acid, taurocholic acid, ethocholicacid, deoxycholic acid, chenodeoxycholic acid, dehydrocholic acid,glycodeoxycholic acid, cyclodextrins and the like in an amount in therange between about 0.2 and 15 weight percent, preferably between about0.5 and 4 weight percent, most preferably about 2 weight percent.

Delivery of the compounds of the present invention to the subject overprolonged periods of time, for example, for periods of one week to oneyear, may be accomplished by a single administration of a controlledrelease system containing sufficient active ingredient for the desiredrelease period. Various controlled release systems, such as monolithicor reservoir-type microcapsules, depot implants, osmotic pumps,vesicles, micelles, liposomes, transdermal patches, iontophoreticdevices and alternative injectable dosage forms may be utilized for thispurpose. Localization at the site to which delivery of the activeingredient is desired is an additional feature of some controlledrelease devices, which may prove beneficial in the treatment of certaindisorders.

One form of controlled release formulation contains the polypeptide orits salt dispersed or encapsulated in a slowly degrading, non-toxic,non-antigenic polymer such as copoly(lactic/glycolic) acid, as describedin the pioneering work of Kent, Lewis, Sanders, and Tice, U.S. Pat. No.4,675,189. The compounds or, preferably, their relatively insolublesalts, may also be formulated in cholesterol or other lipid matrixpellets, or silastomer matrix implants. Additional slow release, depotimplant or injectable formulations will be apparent to the skilledartisan. See, for example, Sustained and Controlled Release DrugDelivery Systems, J. R. Robinson ed., Marcel Dekker, Inc., New York,1978, and R. W. Baker, Controlled Release of Biologically Active Agents,John Wiley & Sons, New York, 1987.

The following specific Examples are intended to illustrate the synthesisand testing of representative compounds of the invention and should notbe construed as limiting the scope of the claims. In the Examples,"m.p." is melting point, " α!_(D) ²⁵ " is the optical activity at 25° C.at the given concentration in the indicated solvent, "FAB" is fast atombombardment mass spectrometry, and "AAA" is amino acid analysis, withexpected values in parentheses following the observed values. The aminoacid analyses were conducted on a Hewlett-Packard AminoQuant Analyzerfollowing the manufacturer's recommended protocols. Primary amino acidswere derivatized with o-phthalaldehyde; secondary amino acids with Fmoc.Fluorescent detection of the derivatized amino acids was used forquantification. The protected amino acids were obtained from AppliedBiosystems Inc. (Foster City, Calif.).

EXAMPLE I

Compound 1 (SEQ ID NO:7) was prepared on 0.5 mmol scale by the solidphase method on 4-methylbenzhydrylamine resin, using an automatedApplied Biosystems Model 430A peptide synthesizer. The α-amino groupswere protected with t-butoxycarbonyl (Boc). The side chain protectinggroups were: benzyl (Bzl) for Asp, Glu, and Ser; tosyl (Tos) for Arg;benzyloxymethyl (Bom) for His; and 2-chlorobenzyl (Cl-z) for Lys. Theamino acids were coupled sequentially usingN,N-dicyclohexylcarbodiimide/1-hydroxybenzotriazole (DCC/HOBt) followingStewart and Young (supra). After each amino acid coupling, the peptidewas acetylated using a mixture of acetic anhydride anddiisopropylethylamine in N-methylpyrrolidone. The completed peptide wascleaved from the resin with simultaneous deprotection of the side chainprotecting groups using anhydrous HF (25 mL) in the presence of anisole(2.5 mL) at -10° C. for 30 minutes and at 0° C. for 60 minutes. Afterevaporation of the HF in vacuo, the residue was washed with anhydrousether, and the crude peptide extracted with 10% acetic acid.Lyophilization of the 10% acetic acid extract gave 900 mgs of crudeproduct. The peptide was purified by medium pressure ODS reversed phasecolumn chromatography using a gradient of 22-45% CH₃ CN in 0.1% TFA. Theproduct eluted in three fractions, which were concentrated andlyophilized to give 130 mgs of white solid of >98% purity.

    __________________________________________________________________________    Compound 1:    AVSEHQLLHDKGKSIQDLRRRELLEKLLEKLHTA-NH.sub.2 (SEQ ID NO:7)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:7)    __________________________________________________________________________    Physical Data: m.p. 150-159° C.    α!.sub.D.sup.25    -34.88° (c 0.16, H.sub.2 O)    FAB (C.sub.175 H.sub.300 N.sub.56 O.sub.51):  M+H!.sup.+  4005.5    AAA:  Asp, 1.9(2);                    Glu, 5.6(6);                              Ser, 1.6(2);                                       His, 2.7(3);          Gly, 1.0(1);                    Thr, 0.9(1);                              Ala, 1.9(2);                                       Arg, 2.8(3)          Val, 1.0(1);                    Ile, 0.9(1);                              Leu, 7.3(8);                                       Lys, 4.0(4).    __________________________________________________________________________    Similarly, Compounds 2, 5-18, 21-27, 29-36, 38-48, 50-54,    58-64 and 66-70 were prepared and characterized,    substituting PAM resin as needed for the synthesis of    hydroxy-terminal polypeptides.    Compound 2:    AVSEHQLLHDKGKSIQDLRRRELLEKLLEKLHTA-OH (SEQ ID NO: 6)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                OH (SEQ ID NO:6)    __________________________________________________________________________    Physical Data: m.p. 154-170° C.    α!.sub.D.sup.25    -49.35° (c 0.46, H.sub.2 O)    FAB (C.sub.175 H.sub.301 N.sub.57 O.sub.50):  M+H!.sup.+  4005.0    AAA:  Asp, 2.1(2);                    Glu, 5.9(6);                              Ser, 1.7(2);                                       His, 2.9(3);          Gly 1.1(1);                    Thr, 1.0(1);                              Ala, 1.9(2);                                       Arg, 3.0(3);          Val, 1.2(1);                    Ile, 1.0(1);                              Leu, 7.8(8);                                       Lys, 4.2(4).    __________________________________________________________________________    Compound 5:    AVSEHQLLHDKGKSIQDLRRRELLERLLERLHTA-OH (SEQ ID NO: 15)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Glu                                              Arg                20             25             30    Leu       His          Thr             Ala                OH (SEQ ID NO:15)    __________________________________________________________________________    Physical Data: m.p. 147-165° C.    α!.sub.D.sup.25    -49.17° (c 0.66, H.sub.2 O)    FAB (C.sub.175 H.sub.299 N.sub.59 O.sub.52):  M+H!.sup.+  4061    AAA:  Asp, 2.1(2);                    Gly, 6.1(6);                              Ser, 1.8(2);                                       His, 3.1(3);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 2.0(2);                                       Arg, 5.0(5);          Val, 1.0(1);                    Ile, 0.9(1);                              Leu, 7.7(8);                                       Lys, 1.9(2).    __________________________________________________________________________    Compound 6:    AVSEHQLLHDRGRSIQDLRRRELLERLLERLHTA-OH (SEQ ID NO:16)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Arg                                     Gly                                        Arg                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Glu                                              Arg                20             25             30    Leu       His          Thr             Ala                OH (SEQ ID NO:16)    __________________________________________________________________________    Physical Data: m.p. 150-170° C.    α!.sub.D.sup.25    -48.65° (c 0.54, H.sub.2 O)    FAB (C.sub.175 H.sub.299 N.sub.63 O.sub.52):  M+H!.sup.+  4118.0    AAA:  Asp, 2.1(2);                    Glu, 6.1(6);                              Ser, 1.8(2);                                       His, 3.2(3);          Gly, 1.2(1);                    Thr, 1.0(1);                              Ala, 2.0(2);                                       Arg, 6.9(7);          Val, 1.0(1);                    Ile, 1.0(1);                              Leu, 7.8(8).    __________________________________________________________________________    Compound 7:    AVSEHQLLHDRGRSIQDLRRRELLERLLKRLHTA-OH (SEQ ID NO:17)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Arg                                     Gly                                        Arg                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Lys                                              Arg                20             25             30    Leu       His          Thr             Ala                OH (SEQ ID NO:17)    __________________________________________________________________________    Physical Data: m.p. 177-182° C.    α!.sub.D.sup.25    -46.17° (c 0.14, H.sub.2 O)    FAB (C.sub.176 H.sub.304 N.sub.64 O.sub.50):  M+H!.sup.+  4117    AAA:  Asp, 2.0(2);                    Glu, 4.8(5);                              Ser, 1.8(2);                                       His, 3.2(3);          Gly, 1.1(1);                    Thr, 0.9(1);                              Ala, 1.9(2);                                       Arg, 6.7(7);          Val, 1.0(1);                    Ile, 1.0(1);                              Lys, 7.7(8);                                       Lys, 0.9(1).    __________________________________________________________________________    Compound 8:    AVSEHQLLHDKGKSIQDLRRRELLEKLLRKLHTA-OH (SEQ ID NO:5)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Arg                                              Lys                20             25             30    Leu       His          Thr             Ala                OH (SEQ ID NO:5)    __________________________________________________________________________    Physical Data: m.p. 147-165° C.    α!.sub.D.sup.25    -49.17° (c 0.66, H.sub.2 O)    FAB (C.sub.176 H.sub.305 N.sub.59 O.sub.49):  M+H!.sup.+  4033.0    AAA:  Asp, 2.0(2);                    Glu, 4.8(5);                              Ser, 1.8(2);                                       His, 2.7(3);          Gly, 1.1(1);                    Thr, 0.9(1);                              Ala, 2.0(2);                                       Arg, 3.9(4);          Val, 1.0(1);                    Ile, 1.0(1);                              Leu, 7.9(8);                                       Lys, 4.0(4).    __________________________________________________________________________    Compound 9:    AVSEHQLLHDKGKSIQDLRRRELLEKLLEKLHTAGRR-OH (SEQ ID NO:10)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                Gly                   Arg                      Arg                         OH (SEQ ID NO:10)                35    __________________________________________________________________________    Physical Data: m.p. 158-160° C.    α!.sub.D.sup.25    -44.76° (c 0.1, H.sub.2 O)    FAB (C.sub.189 H.sub.326 N.sub.64 O.sub.55):  M!.sup.+  4375.0    AAA:  Asp, 2.0(2);                    Glu, 5.9(6);                              Ser, 1.7(2);                                       His, 2.9(3);          Gly, 2.3(2);                    Thr, 1.0(1);                              Ala, 1.9(2);                                       Arg, 5.0(5);          Val, 1.2(1);                    Ile, 1.0(1);                              Leu, 7.8(8);                                       Lys, 4.3(4).    __________________________________________________________________________    Compound 10:    AVSEAQLLHDLGKSIQDLRRRELLEKLLEKLHAL-OH (SEQ ID NO: 14)    Ala       Val          Ser             Glu                Ala                   Gln                      Leu                         Leu                            His                               Asp                                  Leu                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Ala             Leu                OH (SEQ ID NO:14)    __________________________________________________________________________    Physical Data: m.p. 170-175° C.    α!.sub.D.sup.25    -31.59° (c 0.54, H.sub.2 O)    FAB (C.sub.174 H.sub.300 N.sub.52 O.sub.51):  M+H!.sup.+  3936.0    AAA:  Asp, 2.0(2);                    Glu, 6.0(6);                              Ser, 1.8(2);                                       His, 2.0(2);          Gly, 1.2(1);                    Ala, 3.0(3);                              Arg, 2.8(3);                                       Val, 1.1(1);          Ile, 1.0(1);                    Leu, 9.9(10);                              Lys, 3.0(3).    __________________________________________________________________________    Compound 11:    AVSEHQLLHDKGKSIQDLRRRELLEKLLELLKEL-NH.sub.2 (SEQ ID NO:11)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       Lys          Glu             Leu                NH.sub.2 (SEQ ID NO:11)    __________________________________________________________________________    Physical Data: m.p. 172-174° C.    α!.sub.D.sup.25    -43.29° (c 0.2, H.sub.2 O)    FAB (C.sub.179 H.sub.311 N.sub.55 O.sub.52):  M+H!.sup.+  4065.8    AAA:  Asp, 2.2(2);                    Glu, 7.7(7);                              Ser, 1.7(2);                                       His, 2.0(2);          Gly, 1.0(1);                    Ala, 1.0(1);                              Arg, 3.0(3);                                       Val, 1.1(1);          Ile, 1.0(1);                    Leu, 9.3(9);                              Lys, 5.1(5).    __________________________________________________________________________    Compound 12:    AVSEIQFXHNLGKHLSSXERVELLEKLLEKLHNY-NH.sub.2 (X = Nle, SEQ ID NO: 23)    Ala       Val          Ser             Glu                Ile                   Gln                      Phe                         Nle                            His                               Asn                                  Leu                                     Gly                                        Lys                                           His                                              Leu    1           5              10             15    Ser       Ser          Nle             Glu                Arg                   Val                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Asn             Tyr                NH.sub.2 (SEQ ID NO:23)    __________________________________________________________________________    Physical Data: m.p. 178° C.    α!.sub.D.sup.25    -36.88° (c 0.4, H.sub.2 O)    FAB (C.sub.182 H.sub.295 N.sub.50 O.sub.51):  M+H!.sup.+  4001.6    AAA:  Asp, 2.1(2);                    Glu, 6.5(6);                              Ser, 2.7(3);                                       His, 3.1(3)          Gly, 1.1(1);                    Ala, 1.0(1);                              Arg, 1.0(1);                                       Tyr, 0.8(1);          Val, 2.0(2);                    Phe, 1.0(1);                              Ile, 0.9(1);                                       Leu + Nle, 8.5(7 + 2);          Lys, 3.1(3).    __________________________________________________________________________    Compound 13:    AVSEIQFXHNLGKHLSSXRRRELLEKLLEKLHNY-NH.sub.2 (X = Nle, SEQ ID NO: 24)    Ala       Val          Ser             Glu                Ile                   Gln                      Phe                         Nle                            His                               Asn                                  Leu                                     Gly                                        Lys                                           His                                              Leu    1           5              10             15    Ser       Ser          Nle             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Asn             Tyr                NH.sub.2 (SEQ ID NO:24)    __________________________________________________________________________    Physical Data: m.p. 260° C.    α!.sub.D.sup.25    -37.02° (c 0.2, H.sub.2 O)    FAB (C.sub.184 H.sub.304 N.sub.56 O.sub.49):  M+H!.sup.+  4084    AAA:  Asp, 2.1(2);                    Glu, 5.5(5);                              Ser, 2.6(3);                                       His, 3.1(3);          Ala, 1.0(1);                    Gly, 1.1(1);                              Arg, 3.2(3);                                       Tyr, 1.0(1);          Val, 1.0(1);                    Phe, 1.0(1);                              Ile, 1.0(1);                                       Leu, 9.0(9);          Lys, 3.0(3).    __________________________________________________________________________    Compound 14:    AVSEHQLLHDKGKSIQDLRRRALAEALAEALHTA-NH.sub.2 (SEQ ID NO:20)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Ala                         Leu                            Ala                               Glu                                  Ala                                     Leu                                        Ala                                           Glu                                              Ala                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:20)    __________________________________________________________________________    Physical Data: m.p. 190-225° C.    α!.sub.D.sup.25    -56.58° (c 0.36, H.sub.2 O)    FAB (C.sub.161 H.sub.272 N.sub.54 O.sub.49):  M+H!.sup.+  3747.0    AAA:  Asp, 2.1(2);                    Glu, 4.9(5);                              Ser, 1.7(2);                                       His, 2.6(3);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 7.6(7);                                       Arg, 2.8(3)          Val, 1.2(1);                    Ile, 1.0(1);                              Leu, 6.6(6);                                       Lys, 1.9(2).    __________________________________________________________________________    Compound 15:    AVSEHQLLHDKGKSIQDLARRELLEKLLEKLHTA-NH.sub.2 (SEQ ID NO:12)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Ala                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:12)    __________________________________________________________________________    Physical Data: m.p. 170-180° C.    α!.sub.D.sup.25    -48.19° (c 0.2, H.sub.2 O)    FAB (C.sub.172 H.sub.293 N.sub.53 O.sub.51):  M+H!.sup.+  3919.0    AAA:  Asp, 2.1(2);                    Glu, 6.1(6);                              Ser, 1.7(2);                                       His, 3.1(3);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 3.0(3);                                       Arg, 2.1(2);          Val, 1.1(1);                    Ile, 1.0(1);                              Leu, 8.0(8);                                       Lys, 4.4(4).    __________________________________________________________________________    Compound 16:    AVSEHQLLHDKGKSIQDLRRAELLEKLLEKLHTA-NH.sub.2 (SEQ ID NO:13)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Ala                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:13)    __________________________________________________________________________    Physical Data: m.p. 190-195° C.    α!.sub.D.sup.25    -50.50° (c 0.4, H.sub.2 O)    FAB (C.sub.172 H.sub.293 N.sub.53 O.sub.51):  M+H!.sup.+  3919.0    AAA:  Asp, 2.1(2);                    Glu, 6.0(6);                              Ser, 1.8(2);                                       His, 3.1(3);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 3.0(3);                                       Arg, 2.1(2);          Val, 1.1(1);                    Ile, 1.0(1);                              Leu, 7.5(8);                                       Lys, 4.2(4).    __________________________________________________________________________    Compound 17:    AVSEHQLLHDKGKSIQDLRRRSLLSSLLSSLHTA-NH.sub.2 (SEQ ID NO:21)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Ser                         Leu                            Leu                               Ser                                  Ser                                     Leu                                        Leu                                           Ser                                              Ser                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:21)    __________________________________________________________________________    Physical Data: m.p. 195-204° C.    α!.sub.D.sup.25    -67.11° (c 0.3, H.sub.2 O)    FAB (C.sub.163 H.sub.280 N.sub.54 O.sub.50):  M+H!.sup.+  3796.0    AAA:  Asp, 2.1(2);                    Glu, 2.9(3);                              Ser, 6.8(7);                                       His, 3.1(3);          Gly, 1.2(1);                    Thr, 1.0(1);                              Ala, 2.0(2);                                       Arg, 3.0(3);          Val, 1.0(1);                    Ile, 1.0(1);                              Leu, 8.2(8);                                       Lys, 2.0(2).    __________________________________________________________________________    Compound 18:    AVSEHQLLHDKGKSIQDLRRRAFYDKVAEKLHTA-NH.sub.2 (SEQ ID NO:22)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Ala                         Phe                            Tyr                               Asp                                  Lys                                     Val                                        Ala                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:22)    __________________________________________________________________________    Physical Data: m.p. 200-207° C.    α!.sub.D.sup.25    -60.26° (c 0.6, H.sub.2 O)    FAB (C.sub.174 H.sub.284 N.sub.56 O.sub.50):  M+H!.sup.+  3960.0    AAA:  Asp, 2.9(3),                    Glu, 3.5(4);                              Ser, 1.4(2);                                       His, 2.6(3);          Gly, 0.9(1);                    Thr, 1.0(1);                              Ala, 4.0(4);                                       Arg, 3.0(3);          Tyr, 0.9(1);                    Val, 1.9(2);                              Phe, 1.1(1);                                       Ile, 0.9(1);          Leu, 3.6(4);                    Lys, 4.1(4).    __________________________________________________________________________    Compound 21:    AVSEIQFLHN LGKHLSSLRR RELLEKLLEK LHNY-NH.sub.2 (SEQ ID NO:35)    Ala       Val          Ser             Glu                Ile                   Gln                      Phe                         Leu                            His                               Asn                                  Leu                                     Gly                                        Lys                                           His                                              Leu    1           5              10             15    Ser       Ser          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Asn             Tyr                NH.sub.2 (SEQ ID NO:35)    __________________________________________________________________________    Physical Data: m.p. 148-155° C.    α!.sub.D.sup.25   -45.97    (c 0.26, H.sub.2 O)    FAB (C.sub.184 H.sub.304 N.sub.56 O.sub.49):  M+H!.sup.+  4084    AAA:  Asx, 2.1(2);                    Glx, 5.0(5);                              Ser, 2.7(3);                                       His, 3.0(3);          Gly, 1.0(1);                    Ala, 0.9(1);                              Arg, 3.1(3);                                       Tyr, 0.9(1);          Val, 1.0(1);                    Phe, 0.9(1);                              Ile, 0.9(1);                                       Leu, 9.3(9);          Lys, 3.2(3).    __________________________________________________________________________    Compound 24:    AVSEHQLLHD KGKSIQDLKL KELLEKLLEK LHTA-NH.sub.2 (SEQ ID NO:38)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Lys                Leu                   Lys                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:38)    __________________________________________________________________________    Physical Data: m.p. 175-182° C.    α!.sub.D.sup.25   -49.99    (c 0.47, H.sub.2 O)    FAB (C.sub.175 H.sub.299 N.sub.49 O.sub.51):  M+H!.sup.+  3906.5    AAA:  ASX, 2.1(2);                    Glx, 6.5(6);                              Ser, 1.8(2);                                       His, 3.1(3);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 2.1(2);                                       Val, 1.1(1);          Ile, 1.0(1);                    Leu, 9.1(9);                              Lys, 6.5(6).    __________________________________________________________________________    Compound 25:    AVSEHQLLHD KGKSIQDLRR RELLERLLER LHTA-NH.sub.2 (SEQ ID NO:39)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Glu                                              Arg                20             25             30    Leu       His          Thr             Ala--NH.sub.2 (SEQ ID NO:39)    __________________________________________________________________________    Physical Data: m.p. 136.5-153.5° C.    α!.sub.D.sup.25    -32.57 (c 0.13, H.sub.2 O)    FAB (C.sub.175 H.sub.300 N.sub.60 O.sub.51):  M+H!.sup.+  4060.8    AAA:  Asx, 2.2(2);                    Glx, 6.2(6);                              Ser, 1.8(2);                                       His, 3.2(3);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 2.1(2);                                       Arg, 5.2(5);          Val, 1.1(1);                    Ile, 1.1(1);                              Leu, 8.4(8);                                       Lys, 2.2(2).    __________________________________________________________________________    Compound 26:    AVSEHQLLHD KGKSIQDLRR RELLERLLER LHTAP-OH (SEQ ID NO:40)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Glu                                              Arg                20             25             30    Leu       His          Thr             Ala                Pro                   OH (SEQ ID NO:40)    __________________________________________________________________________    Physical Data: m.p. 125.8-127.2° C.    α!.sub.D.sup.25    -54.62 (c 0.23, H.sub.2 O)    FAB (C.sub.180 H.sub.306 N.sub.60 O.sub.53):  M+H!.sup.+  4158.0    AAA:  Asx, 2.1(2);                    Glx, 6.2(6);                              Ser, 1.8(2);                                       His, 2.9(3);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 2.0(2);                                       Arg, 5.1(5);          Val, 1.0(1);                    Ile, 1.0(1);                              Leu, 8.0(8);                                       Lys, 2.1(2);          Pro, 1.1(1).    __________________________________________________________________________    Compound 27:    AVSEHQLLHD KGKSIQDLRR RELLERLLER LHTAGRR-OH (SEQ ID NO:41)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Glu                                              Arg                20             25             30    Leu       His          Thr             Ala                Gly                   Arg                      Arg                         OH (SEQ ID NO:41)                35    __________________________________________________________________________    Physical Data: m.p. 106-137.3° C.    α!.sub.D.sup.25    -39.55 (c 0.67, H.sub.2 O)    FAB (C.sub.189 H.sub.326 N.sub.68 O.sub.55):  M+H!.sup.+  4430.5    AAA:  Asx, 2.1(2);                    Glx, 5.9(6);                              Ser, 1.6(2);                                       His, 2.7(3);          Gly, 2.2(2);                    Thr, 1.0(1);                              Ala, 1.8(2);                                       Arg, 7.3(7);          Val, 0.8(1);                    Ile, 1.0(1);                              Leu, 8.1(8);                                       Lys, 2.1(2).    __________________________________________________________________________    Compound 29:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTY-NH.sub.2 (SEQ ID NO:43)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Tyr                NH.sub.2 (SEQ ID NO:43)    __________________________________________________________________________    Physical Data: m.p. 160-172° C.    α!.sub.D.sup.25   -49.85    (c 0.34, H.sub.2 O)    FAB (C.sub.181 H.sub.304 N.sub.56 O.sub.52):  M+H!.sup.+  4096.9    AAA:  Asx, 2.0(2);                    Glx, 5.6(6);                              Ser, 1.7(2);                                       His, 3.1(3);          Gly, 1.1(1);                    Thr, 0.9(1);                              Ala, 0.9(1);                                       Arg, 3.0(3);          Tyr, 0.9(1);                    Val, 1.0(1);                              Ile, 1.0(1);                                       Leu, 7.7(8);          Lys, 4.4(4).    __________________________________________________________________________    Compound 30:    AVSEHQLLHD KGYSIQDLRR RELLEKLLEK LHTA-NH.sub.2 (SEQ ID NO:44)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Tyr                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:44)    __________________________________________________________________________    Physical Data: m.p. 130-171° C.    α!.sub.D.sup.25   -40.65    (c 0.34, H.sub.2 O)    FAB (C.sub.178 H.sub.297 N.sub.55 O.sub.52):  M+H!.sup.+  4039.4    AAA:  Asx, 2.0(2);                    Glx, 5.5(6);                              Ser, 1.8(2);                                       His, 3.4(3);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 2.0(2);                                       Arg, 2.9(3);          Tyr, 0.8(1);                    Val, 1.0(1);                              Ile, 0.9(1);                                       Leu, 7.9(8);          Lys, 3.4(3).    __________________________________________________________________________    Compound 31:    AVSEHQLLHD KGCSIQDLRR RELLEKLLEK LHTA-NH.sub.2 (SEQ ID NO:45)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Cys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:45)    __________________________________________________________________________    Physical Data: m.p. 140-160° C.    α!.sub.D.sup.25   -44.48    (c 0.25, H.sub.2 O)    FAB (C.sub.172 H.sub.293 N.sub.55 O.sub.51 S.sub.1):  M+H!.sup.+  3979    AAA:  Asx + Cys, 3.0(2 + 1);                    Glx, 5.6.(6);                              Ser, 1.7(2);                                       His, 3.0(3);          Gly, 1.0(1);                    Thr, 0.9(1);                              Ala, 1.9(2);                                       Arg, 2.5(3);          Val, 1.0(1);                    Ile, 0.9(1);                              Leu, 7.5(8);                                       Lys, 3.3(3).    __________________________________________________________________________    Compound 32:    AVSEHQLLHD KGXSIQDLRR RELLEKLLEH LHTA-NH.sub.2 (SEQ ID NO:46)    (X = Cys(CH.sub.2 CONH(CH.sub.2).sub.2 NH(biotinyl)))    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Xaa                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2, (Xaa =    Cys(CH.sub.2 CONH(CH.sub.2).sub.2 NH(biotinyl))), (SEQ ID    __________________________________________________________________________    NO:46)    Physical Data: m.p. (not determined)    α!.sub.D.sup.25   (not    determined)    FAB (C.sub.186 H.sub.316 N.sub.59 O.sub.54 S.sub.2):  M+H!.sup.+  4306.6    AAA:  Asx, 2.2(2);                    Glx, 6.1(6);                              Ser, 1.8(2);                                       His, 3.8(3);          Gly, 1.0(1);                    Thr, 1.0(1);                              Ala, 2.0(2);                                       Arg, 3.1(3);          Val, 1.1(1);                    Ile, 0.9(1);                              Leu, 8.3(3);                                       Lys, 3.3(3).    __________________________________________________________________________    Compound 33:    AVSEHQLLHD KGXSIQDLRR RELLEKLLEK LHTA-NH.sub.2 (SEQ ID NO:47)    (X = Lys(7-dimethylamino-2-oxo-2H-1-benxopyran-4-acetyl))    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Xaa                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2, (Xaa = Lys(7-dimethylamino-2-oxo-2H-1-    benxopyran-4-acetyl), (SEQ ID NO:47)    __________________________________________________________________________    Physical Data: m.p. 135-205° C.    α!.sub.D.sup.25   -26.92    (c 0.104, 50% aq. HOAc)    FAB (C.sub.188 H.sub.311 N.sub.57 O.sub.54):  M+H!.sup.+  4233    AAA:  Asx, 1.9(2);                    Glx, 6.3(6);                              Ser, 1.7(2);                                       His, 3.2(3);          Gly, 1.0(1);                    Thr, 1.1(1);                              Ala, 2.0(2);                                       Arg, 3.2(3);          Val, 1.1(1);                    Ile, 0.9(1);                              Leu, 8.2(8);                                       Lys, 4.5(4).    __________________________________________________________________________    Compound 34:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTAG-OH (SEQ ID NO:48)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                Gly                   OH (SEQ ID NO:48)                35    __________________________________________________________________________    Physical Data: m.p. 92.1-146.6° C.    α!.sub.D.sup.25    -40.76 (c 0.34, H.sub.2 O)    FAB (C.sub.177 H.sub.302 N.sub.56 O.sub.53): (M+H!.sup.+  4062.0    AAA:  Asx, 2.0(2);                    Glx, 5.7(6);                              Ser, 1.8(2);                                       His, 3.0(3);          Gly, 2.2(2);                    Thr, 0.9(1);                              Ala, 1.9(2);                                       Arg, 2.8(3);          Val, 1.2(1);                    Ile, 0.9(1);                              Leu, 7.5(8);                                       Lys, 4.2(4).    __________________________________________________________________________    Compound 35:    AVSX.sub.1 HQLLHX.sub.2 KGKSIQX.sub.2 LRR RX.sub.1 LLX.sub.1 KLLX.sub.1 K    LHA-OH (SEQ ID NO:49)    (X.sub.1 = Glu(OCH.sub.3); X.sub.2 = Asp(OCH.sub.3))    Ala       Val          Ser             Xaa.sub.1                His                   Gln                      Leu                         Leu                            His                               Xaa.sub.2                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Xaa.sub.2          Leu             Arg                Arg                   Arg                      Xaa.sub.1                         Leu                            Leu                               Xaa.sub.1                                  Lys                                     Leu                                        Leu                                           Xaa.sub.1                                              Lys                20             25             30    Leu       His          Ala             OH (Xaa.sub.1 = Glu(OCH.sub.3); Xaa.sub.2 = Asp(OCH.sub.3)) (SEQ             ID NO:49)    __________________________________________________________________________    Physical Data: m.p. (not determined)    α!.sub.D.sup.25   -21.96 (c    0.132, H.sub.2 O)    FAB (C.sub.181 H.sub.311 N.sub.55 O.sub.52):  M+H!.sup.+  4089.0    AAA:  Asx, 2.1(2);                    Glx, 6.3(6);                              Ser, 1.8(2);                                       His, 3.3(3);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 2.0(2);                                       Arg, 3.1(3);          Val, 1.1(1);                    Ile, 0.9(1);                              Leu, 8.0(8);                                       Lys, 4.2(4).    __________________________________________________________________________    Compound 36:    AVSX.sub.1 HOLLHX.sub.2 KGKSIQX.sub.2 LRR RX.sub.1 LLX.sub.1 KLLX.sub.1 K    LHA-OCH.sub.3 (SEQ ID NO:50)    (X.sub.1 = Glu(OCH.sub.3); X.sub.2 = Asp(OCH.sub.3)    Ala       Val          Ser             Xaa.sub.1                His                   Gln                      Leu                         Leu                            His                               Xaa.sub.2                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Xaa.sub.2          Leu             Arg                Arg                   Arg                      Xaa.sub.1                         Leu                            Leu                               Xaa.sub.1                                  Lys                                     Leu                                        Leu                                           Xaa.sub.1                                              Lys                20             25             30    Leu       His          Ala             OCH.sub.3 (Xaa.sub.1  = Glu(OCH.sub.3); Xaa.sub.2             = Asp(OCH.sub.3)) (SEQ ID NO:50)    __________________________________________________________________________    Physical Data: m.p. (not determined)    α!.sub.D.sup.25   -46.80 (c    0.07, H.sub.2 O)    FAB (C.sub.182 H.sub.313 N.sub.55 O.sub.52):  M+H!.sup.+  4103    AAA:  Asx, 2.1(2);                    Glx, 6.2(6);                              Ser, 1.4(2);                                       His, 3.0(3);          Gly, 1.1(1);                    Thr, 1.1(1);                              Ala, 1.7(2);                                       Arg, 3.2(3);          Val, 0.6(1);                    Ile, 0.9(1);                              Leu, 8.0(8);                                       Lys, 4.1(4).    __________________________________________________________________________    Compound 38:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTAP-OH (SEQ ID NO:52)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                Pro                   OH (SEQ ID NO:52)                35    __________________________________________________________________________    Physical Data: m.p. 152.1-186.5° C.    α!.sub.D.sup.25    -55.91 (c 0.33, H.sub.2 O)    FAB (C.sub.180 H.sub.306 N.sub.56 O.sub.53):  M+H!.sup.+  4102.6    AAA:  Asx, 2.0(2);                    Glx, 5.6(6);                              Ser, 1.7(2);                                       His, 2.9(3);          Gly, 1.1(1);                    Thr, 0.9(1);                              Ala, 1.9(2);                                       Arg, 2.9(3);          Val, 1.2(1);                    Ile, 1.0(1);                              Leu, 7.7(8);                                       Lys, 4.3(4).    __________________________________________________________________________    Compound 39:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTP-OH (SEQ ID NO:53)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Pro                OH (SEQ ID NO:53)    __________________________________________________________________________    Physical Data: m.p. 120-148.2° C.    α!.sub.D.sup.25    -52.78 (c 0.47, H.sub.2 O)    FAB (C.sub.177 H.sub.301 N.sub.55 O.sub.52):  M+H!.sup.+  4031.0    AAA:  Asx, 2.0(2);                    Glx, 5.5(6);                              Ser, 1.8(2);                                       His, 2.9(3);          Gly, 1.0(1);                    Thr, 1.0(1);                              Ala, 0.9(1);                                       Arg, 2.9(3);          Val, 1.2(1);                    Ile, 0.9(1);                              Leu, 7.5(8);                                       Lys, 3.6(3);          Pro, 0.9(1).    __________________________________________________________________________    Compound 40:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTP-NH.sub.2 (SEQ ID NO:54)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Pro                NH.sub.2 (SEQ ID NO:54)    __________________________________________________________________________    Physical Data: m.p. 133.9-155.1° C.    α!.sub.D.sup.25    -54.22 (c 0.37, H.sub.2 O)    FAB (C.sub.177 H.sub.302 N.sub.56 O.sub.51):  M+H!.sup.+  4030.7    AAA:  Asx, 2.0(2);                    Glx, 5.6(6);                              Ser, 1.9(2);                                       His, 2.9(3);          Gly, 1.1(1);                    Thr, 0.9(1);                              Ala, 0.9(1);                                       Arg, 2.8(3);          Val, 1.2(1);                    Ile, 1.1(1);                              Leu, 7.8(8);                                       Lys, 4.2(4).          Pro, 0.9(1).    __________________________________________________________________________    Compound 41:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHP-NH.sub.2 (SEQ ID NO:55)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Pro             NH.sub.2 (SEQ ID NO:55)    __________________________________________________________________________    PhysicaI Data: m.p. 142.8-166.1° C.    α!.sub.D.sup.25    -53.80 (c 0.38, H.sub.2 O)    FAB (C.sub.173 H.sub.295 N.sub.55 O.sub.49):  M+H!.sup.+  3929    AAA:  Asx, 2.0(2);                    Glx, 5.7(6);                              Ser, 1.8(2);                                       His, 3.0(3);          Gly, 1.1(1);                    Ala, 0.9(1);                              Arg, 2.8(3);                                       Val, 1.2(1);          Ile, 0.9(1);                    Leu, 7.4(8);                              Lys, 4.4(4);                                       Pro, 0.9(1).    __________________________________________________________________________    Compound 42:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LP-NH.sub.2 (SEQ ID NO:56)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       Pro          NH.sub.2 (SEQ ID NO:56)    __________________________________________________________________________    Physical Data: m.p. 161.0-177.0° C.    α!.sub.D.sup.25    -61.97 (c 0.19, H.sub.2 O)    FAB (C.sub.167 H.sub.288 N.sub.52 O.sub.48):  M+H!.sup.+  3792.0    AAA:  Asx, 2.2(2);                    Glx, 5.9(6);                              Ser, 1.9(2);                                       His, 2.1(2);          Gly, 1.1(1);                    Ala, 1.0(1);                              Arg, 3.0(3);                                       Val, 1.1(1);          Ile, 1.0(1);                    Leu, 7.9(8);                              Lys, 4.3(4);                                       Pro, 0.9(1).    __________________________________________________________________________    Compound 43:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTRSAW-OH (SEQ ID NO:57)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Arg                Ser                   Ala                      Trp                         OH (SEQ ID NO:57)                35    __________________________________________________________________________    Physical Data: m.p. 181-202° C.    α!.sub.D.sup.25   -45.14    (c 0.19, H.sub.2 O)    FAB (C.sub.195 H.sub.326 N.sub.62 O.sub.56):  M+H!.sup.+  4435.2    AAA:  Asx, 2.0(2);                    Glx, 5.8(6);                              Ser, 2.8(3);                                       His, 2.8(3);          Gly, 1.1(1);                    Thr, 0.9(1);                              Ala, 1.9(2);                                       Arg, 3.7(4);          Ile, 0.9(1);                    Leu, 7.5(8);                              Lys, 4.3(4);                                       Trp, 0.9(1).    __________________________________________________________________________    Compound 44:    AVSEHQLLHD RGRSIQDLRR RELLERLLER LHTAGRRTRSAW-OH (SEQ ID NO:58)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Arg                                     Gly                                        Arg                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Glu                                              Arg                20             25             30    Leu       His          Thr             Arg                Gly                   Arg                      Arg                         Thr                            Arg                               Ser                                  Ala                                     Trp OH (SEQ ID NO:58)                35             40    __________________________________________________________________________    Physical Data: m.p. 130-132.2° C.    α!.sub.D.sup.25    -46.66 (c 0.195, H.sub.2 O)    FAB (C.sub.216 H.sub.365 N.sub.81 O.sub.62):  M+H!.sup.+  5088.8    AAA:  Asx, 2.2(2);                    Glx, 6.0(6);                              Ser, 2.7(3);                                       His, 3.0(3);          Gly, 2.2(2);                    Thr, 2.1(2);                              Ala, 3.0(3);                                       Arg, 10.5(10);          Val, 0.9(1);                    Ile, 1.0(1);                              Leu, 8.2(8);                                       Trp, 1.0(1).    __________________________________________________________________________    Compound 45:    AVSEHQLLHD RGRSIQDLRR RELLERLLER LHTAGRRTRSAW-NH.sub.2 (SEQ ID NO:59)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Arg                                     Gly                                        Arg                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Glu                                              Arg                20             25             30    Leu       His          Thr             Arg                Gly                   Arg                      Arg                         Thr                            Arg                               Ser                                  Ala                                     Trp NH.sub.2 (SEQ ID NO:59)                35             40    __________________________________________________________________________    Physical Data: m.p. 158-174° C.    α!.sub.D.sup.25   -43.57    (c 0.53, H.sub.2 O)    FAB (C.sub.216 H.sub.366 N.sub.82 O.sub.61):  M+H!.sup.+  5087.4    AAA:  Asx, 1.9(2);                    Glx, 5.6(6);                              Ser, 2.6(3);                                       His, 3.3(3);          Gly, 2.1(2);                    Thr, 2.0(2);                              Ala, 2.9(3);                                       Arg, 10.1(10);          Val, 0.9(1);                    Ile, 1.0(1);                              Leu, 8.3(8);                                       Trp 1.1(1).    __________________________________________________________________________    Compound 46:    AVSEHQLLHD RGXSIQDLRR RELLERLLER LHTAGRRTRSAW-OH (SEQ ID NO:60)    (X = Lys(dihydrocinnamoyl))    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Arg                                     Gly                                        Xaa                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Glu                                              Arg                20             25             30    Leu       His          Thr             Arg                Gly                   Arg                      Arg                         Thr                            Arg                               Ser                                  Ala                                     Trp. OH (Xaa =                35             40    Lys(dihydrocinnamoyl), (SEQ ID NO:60)    __________________________________________________________________________    Physical Data: m.p. 165.4-175.2° C.    α!.sub.D.sup.25    -40.43 (c 0.20, H.sub.2 O)    FAB (C.sub.225 H.sub.374 N.sub.80 O.sub.62):  M+H!.sup.+  5191    AAA:  Asx, 2.1(2),                    Glx, 6.3(6);                              Ser, 2.8(3);                                       His, 3.2(3);          Gly, 2.1(2),                    Thr, 2.0(2);                              Ala, 3.2(3);                                       Arg, 9.9(9);          Val, 1.0(1),                    Ile, 0.9(1);                              Leu, 8.6(8);                                       Lys, 1.1(1);          Trp, 1.1(1).    __________________________________________________________________________    Compound 47:    AVSEIQFXHN LGKHLSSXTR SAWLRKKLQD VHNY-NH.sub.2 (SEQ ID NO:61)    (X = norleucine)    Ala       Val          Ser             Glu                Ile                   Gln                      Phe                         Nle                            His                               Asn                                  Leu                                     Gly                                        Lys                                           His                                              Leu    1           5              10             15    Ser       Ser          Nle             Thr                Arg                   Ser                      Ala                         Trp                            Leu                               Arg                                  Lys                                     Lys                                        Leu                                           Gln                                              Asp                20             25             30    Val       His          Asp             Tyr                NH.sub.2 (SEQ ID NO:61)    __________________________________________________________________________    Physical Data: m.p. 140-160° C.    α!.sub.D.sup.25   -56.88    (c 0.16, H.sub.2 O)    FAB (C.sub.180 H.sub.287 N.sub.55 O.sub.58):  M+H!.sup.+  3989.8    AAA:  Asx, 3.0(3);                    Glx, 2.9(3);                              Ser, 3.7(4);                                       His, 2.8(3);          Gly, 1.1(1);                    Thr, 0.9(1);                              Ala, 1.9(2);                                       Arg, 2.0(2);          Tyr, 1.0(1);                    Val, 1.7(2);                              Phe, 0.9(1);                                       Ile, 0.9(1);          Leu + Nle 5.8(2 + 4);                    Lys, 3.4(3);                              Trp, 1.1(1).    __________________________________________________________________________    Compound 48:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTMA-NH.sub.2 (SEQ ID NO:62)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Met                Ala                   NH.sub.2 (SEQ ID NO:62)                35    __________________________________________________________________________    Physical Data: m.p. 140-210° C.    α!.sub.D.sup.25   -47.75    (c 0.178, H.sub.2 O)    FAB (C.sub.180 H.sub.309 N.sub.57 O.sub.52 S.sub.1):  M+H!.sup.+  4135.0    AAA:  Asx, 2.3(2);                    Glx, 6.6(6);                              Ser, 1.4(2);                                       His, 3.2(3);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 2.0(2);                                       Arg, 3.1(3);          Val, 0.9(1);                    Met, 1.1(1);                              Ile, 1.0(1);                                       Leu, 8.8(8);          Lys, 4.4(4).    __________________________________________________________________________    Compound 50:    AVSEHQLLHD KGKSIQDLRR RFFLEKLLEK LHTA-NH.sub.2 (SEQ ID NO:64)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Phe                         Phe                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:64)                35    __________________________________________________________________________    Physical Data: m.p. 136.5-156.8° C.    α!.sub.D.sup.25    -49.89 (c 0.24, H.sub.2 O)    FAB (C.sub.182 H.sub.300 N.sub.56 O.sub.49):  M+H!.sup.+  4056.0    AAA:  Asx, 2.2(2);                    Glx, 5.0(5);                              Ser, 1.9(2);                                       His, 3.3(3);          Gly, 1.0(1);                    Thr, 1.0(1);                              Ala, 2.1(2);                                       Arg, 3.1(3);          Val, 1.0(1);                    Phe, 2.0(2);                              Ile, 0.9(1);                                       Leu, 7.2(7);          Lys, 3.5(4).    __________________________________________________________________________    Compound 51:    AVSEHQLLHD KGKSIQDLRR RELLHKLLEK LHTA-NH.sub.2 (SEQ ID NO:65)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               His                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:65)    __________________________________________________________________________    Physical Data: m.p. 80.7-141.0° C.    α!.sub.D.sup.25    -55.38 (c 0.23, H.sub.2 O)    FAB (C.sub.176 H.sub.300 N.sub.58 O.sub.49):  M+H!.sup.+  4012.8    AAA:  Asx, 2.2(2);                    Glx, 4.9(5);                              Ser, 1.8(2);                                       His, 4.3(4);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 2.0(2);                                       Arg, 3.1(3);          Val, 1.1(1);                    Ile, 1.0(1);                              Leu, 8.1(8);                                       Lys, 3.9(4).    __________________________________________________________________________    Compound 52:    AVSEHQLLHD KGKSIQDLRR RELLEHLLEK LHTA-NH.sub.2 (SEQ ID NO:66)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  His                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:66)    __________________________________________________________________________    Physical Data: m.p. 134.3-157.9° C.    α!.sub.D.sup.25    -50.72 (c 0.45, H.sub.2 O)    FAB (C.sub.175 H.sub.295 N.sub.57 O.sub.51):  M+H!.sup.+  4012.8    AAA:  Asx, 2.1(2);                    Glx, 5.9(6);                              Ser, 1.8(2);                                       His, 4.2(4);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 2.0(2);                                       Arg, 3.0(3);          Val, 1.1(1);                    Ile, 0.9(1);                              Leu, 8.1(8);                                       Lys, 3.1(3).    __________________________________________________________________________    Compound 53:    AVSEHQLLHD KGKSIQDLRR RELLEKLIAK LHTA-NH.sub.2 (SEQ ID NO:67)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Ile                                           Ala                                              Lys                20             25             30    Leu       His          Thr             Ala                NH.sub.2 (SEQ ID NO:67)    __________________________________________________________________________    Physical Data: m.p. 142.7-159.8° C.    α!.sub.D.sup.25    -54.01 (c 0.21, H.sub.2 O)    FAB (C.sub.173 H.sub.295 N.sub.56 O.sub.49):  M+H!.sup.+  3946.0    AAA:  Asx, 2.2(2);                    Glx, 4.9(5);                              Ser, 1.8(2);                                       His, 3.1(3);          Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 3.1(3);                                       Arg, 3.1(3);          Val, 1.0(1);                    Ile, 1.9(2);                              Leu, 7.0(7);                                       Lys, 4.3(4).    __________________________________________________________________________    Compound 54:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEE LHTA-NH.sub.2 (SEQ ID NO:68)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Glu                20             25             30    Ile       His          Thr             Ala                NH.sub.2 (SEQ ID NO:68)    __________________________________________________________________________    Physical Data: m.p. 138-185° C.    α!.sub.D.sup.25   -50.17    (c 0.14, H.sub.2 O)    FAB (C.sub.174 H.sub.295 N.sub.55 O.sub.53):  M+H!.sup.+  4005    AAA:  Asx, 2.2(2);                    Glx, 7.1(7);                              Ser, 1.7(2);                                       His; 2.8(3);          Gly, 1.0(1);                    Thr, 1.0(1);                              Ala, 2.1(2);                                       Arg, 3.1(3);          Val, 1.1(1);                    Ile, 1.7(2);                              Leu, 7.1(7);                                       Lys, 2.7(3).    __________________________________________________________________________    Compound 58:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTRSAWNH.sub.2 (SEQ ID NO:72)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Arg                Ser                   Ala                      Trp                         NH.sub.2 (SEQ ID NO:72)                35    __________________________________________________________________________    Physical Data: m.p. 158-163° C.    α!.sub.D.sup.25   -46.06    (c 0.17, H.sub.2 O)    FAB (C.sub.195 H.sub.327 N.sub.63 O.sub.55):  M+H!.sup.+  4434.8    AAA:  Asx, 2.0(2);                    Glx, 5.5(6);                              Ser, 2.7(3);                                       His, 3.1(3);          Gly, 1.0(1);                    Ala, 1.8(2);                              Arg, 4.0(4);                                       Thr, 0.9(1);          Val, 0.9(1);                    Ile, 0.9(1);                              Leu, 7.5(8);                                       Lys, 3.9(4);          Trp, 1.0(1).    __________________________________________________________________________    Compound 59:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTRSAX-OH (SEQ ID NO:73)    (X = Nal(2) = 3-(2-naphthyl)-L-alanine)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Arg                Ser                   Ala                      3-(2-naphthly)-L-alanine-OH (SEQ ID NO:73)                35    __________________________________________________________________________    Physical Data: m.p. 156-162° C.    α!.sub.D.sup.25   -44.44    (c 0.189, H.sub.2 O)    FAB (C.sub.197 H.sub.328 N.sub.62 O.sub.55):  M+H!.sup.+  4445.6    AAA:  Asx, 2.1(2);                    Glx, 5.5(6);                              Ser, 2.8(3);                                       His, 2.9(3);          Gly, 1.0(1);                    Ala, 2.0(2);                              Arg, 4.0(4);                                       Thr, 0.9(1);          Val, 1.0(1);                    Ile, 0.9(1);                              Leu, 7.5(8);                                       Lys, 4.2(4);          Nal, 1.1(1).    __________________________________________________________________________    Compound 60:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTASAW-OH (SEQ ID NO:74)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                Ser                   Ala                      Trp                         OH (SEQ ID NO:74)                35    __________________________________________________________________________    Physical Data: m.p. 159-164° C.    α!.sub.D.sup.25   -50.94    (c 0.29, H.sub.2 O)    FAB (C.sub.192 H.sub.320 N.sub.60 O.sub.55):  M+H!.sup.+  4349.0    AAA:  Asx, 2.0(2);                    Glx, 5.6(6);                              Ser, 2.7(3);                                       His, 3.2(3);          Gly, 1.0(1);                    Ala, 3.1(3);                              Arg, 2.8(3);                                       Thr, 1.0(1);          Val, 1.1(1);                    Ile, 0.9(1);                              Leu, 7.6(8);                                       Lys, 4.0(4);          Trp, 1.0(1).    __________________________________________________________________________    Compound 61:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTAEIRA-OH (SEQ ID NO:75)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                Glu                   Ile                      Arg                         Ala                            OH (SEQ ID NO.:75)                35    __________________________________________________________________________    Physical Data: m.p. 155-210° C.    α!.sub.D.sup.25   -46.15    (c 0.12, H.sub.2 O)    FAB (C.sub.195 H.sub.334 N.sub.62 O.sub.58):  M+H!.sup.+  4475.8    AAA:  Asx, 2.2(2);                    Glx, 6.9(7);                              Ser, 1.7(2);                                       His, 3.2(3);          Gly, 1.1(1);                    Ala, 3.1(3);                              Arg, 4.0(4);                                       Thr, 0.9(1);          Val, 1.1(1);                    Ile, 1.9(2);                              Leu, 8.1(8);                                       Lys, 4.1(4).    __________________________________________________________________________    Compound 62:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTAEIR-OH (SEQ ID NO:76)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                Glu                   Ile                      Arg                         OH (SEQ ID NO:76)                35    __________________________________________________________________________    Physical Data: m.p. 186-218° C.    α!.sub.D.sup.25   -52.73    (c 0.265, H.sub.2 O)    FAB (C.sub.192 H.sub.329 N.sub.61 O.sub.57):  M+H!.sup.+  4404.4    AAA:  Asx, 2.0(2);                    Glx, 6.6(7);                              Ser, 1.9(2);                                       His, 3.4(3);          Gly, 1.1(1);                    Ala, 2.0(2);                              Arg, 3.8(4);                                       Thr, 1.0(1);          Val, 1.1(1);                    Ile, 1.7(2);                              Leu, 7.9(8);                                       Lys, 4.0(4).    __________________________________________________________________________    Compound 63:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTAEI-OH (SEQ ID. NO:77)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                Glu                   Ile                      OH (SEQ ID NO:77)                35    __________________________________________________________________________    Physical Data: m.p. 169-205° C.    α!.sub.D.sup.25   -50.78    (c 0.51, H.sub.2 O)    FAB (C.sub.186 H.sub.317 N.sub.57 O.sub.56):  M+H!.sup.+  4248.0    AAA:  Asx, 2.2(2);                    Glx, 6.8(7);                              Ser, 1.8(2);                                       His, 3.3(3);          Gly, 1.0(1);                    Ala, 2.0(2);                              Arg, 3.0(3);                                       Thr, 1.0(1);          Val, 1.0(1);                    Ile, 1.8(2);                              Leu, 7.8(8);                                       Lys, 3.6(4).    __________________________________________________________________________    Compound 64:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTAE-OH (SEQ ID NO:78)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             Ala                Glu                   OH (SEQ ID NO:78)                35    __________________________________________________________________________    Physical Data: m.p. 199-205° C.    α!.sub.D.sup.25   -52.47    (c 0.41, H.sub.2 O)    FAB (C.sub.180 H.sub.306 N.sub.56 O.sub.55):  M+H!.sup.+  4135.0    AAA:  Asx, 2.0(2);                    Glx, 6.6(7);                              Ser, 1.9(2);                                       His, 3.3(3);          Gly, 1.1(1);                    Ala, 2.0(2);                              Arg, 2.9(3);                                       Thr, 1.0(1);          Val, 1.1(1);                    Ile, 1.0(1);                              Leu, 8.2(8);                                       Lys, 3.8(4).    __________________________________________________________________________    Compound 66:    SEHQLLHD KGKSIQDLRR RELLEKLLEK LHTA-NH.sub.2 (SEQ ID NO:80)    Ser       Glu          His             Gln                Leu                   Leu                      His                         Asp                            Lys                               Gly                                  Lys                                     Ser                                        Ile                                           Gln                                              Asp    1           5              10             15    Leu       Arg          Arg             Arg                Glu                   Leu                      Leu                         Glu                            Lys                               Leu                                  Leu                                     Glu                                        Lys                                           Leu                                              His                20             25             30    Thr       Ala          NH.sub.2 (SEQ ID NO:80)    __________________________________________________________________________    Physical Data: m.p. 134.2° C.    α!.sub.D.sup.25   -48.12 (c    0.36, H.sub.2 O)    FAB (C.sub.167 H.sub.286 N.sub.54 O.sub.49):  M+H!.sup.+  3834.4    AAA:  Asx, 2.0(2);                    Glx, 5.7(6);                              Ser, 1.7(2);                                       His, 2.9(3);          Gly, 1.0(1);                    Thr, 0.9(1);                              Ala, 1.0(1);                                       Arg, 2.8(3);          Ile, 0.9(1);                    Leu, 7.4(8);                              Lys, 4.3(4).    __________________________________________________________________________    Compound 67:    LLHD KGKSIQDLRR RELLEKLLEK LHTA-NH.sub.2 (SEQ ID NO:81)    Leu       Leu          His             Asp                Lys                   Gly                      Lys                         Ser                            Ile                               Gln                                  Asp                                     Leu                                        Arg                                           Arg                                              Arg    1           5              10             15    Glu       Leu          Leu             Glu                Lys                   Leu                      Leu                         Glu                            Lys                               Leu                                  His                                     Thr                                        Ala                                           NH.sub.2 (SEQ    ID NO:81)                20             25    __________________________________________________________________________    Physical Data: m.p. 128.5-184.5° C.    α!.sub.D.sup.25    -6.53 (c 0.69, MeOH)    FAB (C.sub.148 H.sub.259 N.sub.47 O.sub.41):  M+H!.sup.+  3353    AAA:  Asx, 2.0(2);                    Glx, 4.1(4);                              Ser, 0.9(1);                                       His, 2.1(2);          Gly, 1.0(1);                    Thr, 0.9(1);                              Ala, 1.0(1);                                       Arg, 3.0(3);          Ile, 1.0(1);                    Leu, 8.1(8);                              Lys, 4.2(4).    __________________________________________________________________________    Compound 68:    LHD KGKSIQDLRR RELLEKLLEK LHTA-NH.sub.2 (SEQ ID NO:82)    Leu       His          Asp             Lys                Gly                   Lys                      Ser                         Ile                            Gln                               Asp                                  Leu                                     Arg                                        Arg                                           Arg                                              Glu    1           5              10             15    Leu       Leu          Glu             Lys                Leu                   Leu                      Glu                         Lys                            Leu                               His                                  Thr                                     Ala                                        NH2 (SEQ ID    NO:82)                20             25    __________________________________________________________________________    Physical Data: m.p. 165-210° C.    α!.sub.D.sup.25   -36.05    (c 0.12, H.sub.2 O)    FAB (C.sub.142 H.sub.248 N.sub.46 O.sub.40):  M+H!.sup.+  3239.0    AAA:  Asx, 2.0(2);                    Glx, 3.9(4);                              Ser, 0.9(1);                                       His, 1.9(2);          Gly, 1.0(1);                    Thr, 1.0(1);                              Ala, 1.0(1);                                       Arg, 2.9(3);          Ile, 0.9(1);                    Leu, 6.8(7);                              Lys, 4.2(4).    __________________________________________________________________________    Compound 69:    SEHQLLHD RGRSIQDLRR RELLERLLER LHAGRRTRSAW-OH (SEQ ID NO:83)    Ser       Glu          His             Gln                Leu                   Leu                      His                         Asp                            Arg                               Gly                                  Arg                                     Ser                                        Ile                                           Gln                                              Asp    1           5              10             15    Leu       Arg          Arg             Arg                Glu                   Leu                      Leu                         Glu                            Arg                               Leu                                  Leu                                     Glu                                        Arg                                           Leu                                              His                20             25             30    Leu       His          Arg             Gly                Arg                   Arg                      Thr                         Arg                            Ser                               Ala                                  Trp                                     OH (SEQ ID NO: 83)                35             40    Physical Data: m.p. 150-210° C.    α!.sub.D.sup.25   -18.0    (c 0.64, H.sub.2 O)    __________________________________________________________________________    FAB (C.sub.208 H.sub.351 N.sub.79 O.sub.60):  M+H!.sup.+  4918.6    AAA:  Asx, 2.2(2);                    Glx, 6.2(6);                              Ser, 2.8(3);                                       His, 3.1(3);          Gly, 2.2(2);                    Thr, 2.2(2);                              Ala, 2.2(2);                                       Arg, 10.4(10);          Ile, 1.0(1);                    Leu, 8.0(8);                              Trp, 1.1(1).    __________________________________________________________________________    Compound 70:    LLHD RGRSIQDLRR RELLERLLER LHAGRRTRSAW-OH (SEQ ID NO:84)    Leu       Leu          His             Asp                Arg                   Gly                      Arg                         Ser                            Ile                               Gln                                  Asp                                     Leu                                        Arg                                           Arg                                              Arg    1           5              10             15    Glu       Leu          Leu             Glu                Arg                   Leu                      Leu                         Glu                            Arg                               Leu                                  His                                     Leu                                        His                                           Arg                                              Gly                20             25             30    Arg       Arg          Thr             Arg                Ser                   Ala                      Trp                         OH (SEQ ID NO:84)                35    __________________________________________________________________________    Physical Data: m.p. 150-210° C.    α!.sub.D.sup.25   -41.70    (c 0.36, H.sub.2 O)    FAB (C.sub.189 H.sub.324 N.sub.72 O.sub.52):  M+H!.sup.+  4437.14    AAA:  Asx, 2.1(2);                    Glx, 4.1(4);                              Ser, 1.9(2);                                       His, 2.0(2);          Gly, 2.1(2);                    Thr, 2.0(2);                              Ala, 2.1(2);                                       Arg, 9.7(10);          Ile, 0.9(1);                    Leu, 7.4(8).    __________________________________________________________________________

The side chain cyclized analog (Compound 57) was synthesized as aboveexcept the N.sup.α -Boc-N.sup.ε -Fmoc-Lys and N.sup.α -Boc-N.sup.γ-Fmoc-Asp were placed in positions 13 and 17, respectively. Uponcompletion of the Boc amino acid synthesis, the resin was treated with20% piperidine in DMF at room temperature for 30 minutes. The resin wasfiltered and washed with DMF, MeOH and CH₂ Cl₂. The resin (1.1 g) wassuspended in 10 ml DMF containing 250 mg PyBOP. The pH was adjusted to8-9 with DIEA and the resin stirred for 1 hour. The resin was filtered,washed with DMF and CH₂ Cl₂, then resuspended in DMF. The coupling wasrepeated using 125 mg of PyBOP. The resin was filtered, washed with DMF,MeOH, and CH₂ Cl₂ and dried. The resin was then treated with HF and thepeptide purified as mentioned above. ##STR13##

EXAMPLE II

Met³⁴, Ala³⁵ ! Compound 1, AVSEHQLLHDKGKSIQDLRRRELLEKLLEKLHTMA-NH₂, (SEQID NO:25), was prepared and purified following the procedures of ExampleI. This polypeptide was converted to the homoserine lactone as follows.The purified peptide (160 mgs) was dissolved in 44% formic acid (4 mL).This solution was combined with a premixed solution of cyanogen bromide(700 mgs) and phenol (1.6 mgs) in 44% formic acid (4 mL) at 0° C. Thesolution was stirred at 0° C. for 2 hr and at room temperature for 2hrs. The formation of the product was monitored by HPLC (Vydac® C-18,300 Å, 4.6×250 mm, flow of 1.2 mL/min, gradient 25-45% acetonitrile in0.1% TFA over 10 min). The reaction was complete within 4 hr. Half ofthe sample was concentrated and purified by preparative RP-HPLC (Vydac®C-18, gradient 25-45% acetonitrile in 0.1% TFA). The homoserine lactonepeptide fractions were pooled and lyophilized to give 28 mgs of whitesolid of >95% purity, Compound 4.

    __________________________________________________________________________    Compound 4    AVSEHQLLHDKGKSIQDLRRRELLEKLLEKLHTX (X = hSerlac. SEQ ID    __________________________________________________________________________    NO:9)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             hSerlac (SEQ ID NO:9)    __________________________________________________________________________    Physical Data: m.p. 138-142° C.    α!.sub.D.sup.25    -50.66° (c 0.1, H.sub.2 O)    FAB (C.sub.176 H.sub.299 N.sub.55 O.sub.52):  M+H!.sup.+  4017.61    AAA:   Asp, 2.1(2);                    Glu, 6.1(6);                              Ser, 1.8(2);                                        His, 3.0(3);           Thi, 1.1(1);                    Ala, 1.1(1);                              Arg, 2.7(3);                                        Val, 1.0(1);           Ile, 1.0(1);                    Leu, 8.2(8);                              Lys, 3.8(4);                                        Gly 1.09(1);           hSer, 1.09(1).    __________________________________________________________________________

Similarly, Compound 65 was prepared in accordance with this procedure.

    __________________________________________________________________________    Compound 65:    AVSEIQFX.sub.1 HN KGKHLSSX.sub.1 ER VEWLRKKLQD VHNX.sub.2 (SEQ IF NO:79)    (X.sub.1 = L-norleucine; X.sub.2 = homoserine lactone)    __________________________________________________________________________    Ala       Val          Ser             Glu                Ile                   Gln                      Phe                         Nle                            His                               Asn                                  Lys                                     Gly                                        Lys                                           His                                              Leu    1           5              10             15    Ser       Ser          Nle             Glu                Arg                   Val                      Glu                         Trp                            Leu                               Arg                                  Lys                                     Lys                                        Leu                                           Gln                                              Asp                20             25             30    Val       His          Asn             hSerlac (SEQ ID NO:79)    __________________________________________________________________________    Physical Data: m.p. 166-176° C.    α!.sub.D.sup.25   -52.22    (c 0.25, H.sub.2 O)    FAB (C.sub.180 H.sub.288 N.sub.54 O.sub.50):  M+H!.sup.+ 4008.6    AAA:  Asx, 3.1(3);                   Glx, 4.8(5);                            Ser, 2.9(3);                                        His, 2.9(3);          Gly, 1.1(1);                   Ala, 1.1(1);                            Arg, 2.0(2);                                        Val, 2.7(3);          Phe, 1.0(1);                   Ile, 1.0(1);                            Leu + Nle 5.9 (4 + 2);                                        Lys, 2.8(3).    __________________________________________________________________________

EXAMPLE III

To prepare the homoserine amide, the crude hSerlactone analog, Compound4, was concentrated and treated with 25 mL saturated NH₃ in methanol.The solution was stirred at 0° C. for 2 hr and at room temperature for16 hr. The reaction was monitored by HPLC (Vydace C-18, 300 Å, 4.6×250mm, flow of 1.2 mL/min, gradient 20-45% acetonitrile in 0.1% TFA) andwas complete within 18 hr. The solution was concentrated and purified bypreparative RP-HPLC (Vydac® C-18, gradient of 25-45% acetonitrile in0.1% TFA). The homoserine amide peptide fractions were pooled andlyophilized to give 30 mgs of white solid of >98% purity, Compound 3.

    __________________________________________________________________________    Compound 3    AVSEHQLLHDKGKSIQDLRRRELLEKLLEKLHTXZ-NH.sub.2 (X = hSer, SEQ ID    __________________________________________________________________________    NO:8)    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             hSer NH.sub.2 (SEQ ID NO:8).    __________________________________________________________________________    Physical Data: m.p. 138-142° C.    α!.sub.D.sup.25    -45.97° (c 0.25, H.sub.2 O)    FAB (C.sub.176 H.sub.302 N.sub.56 O.sub.52):  M+H!.sup.+  4033.9    AAA:   Asp, 2.1(2);                    Glu, 6.1(6);                              Ser, 1.6(2);                                        His, 2.8.(3);           Gly, 0.97(1);                    hSer, 0.97(1);                              Thi, 1.0(1);                                        Ala, 1.0(1);           Arg, 2.9(3);                    Val, 1.0(1);                              Ile, 1.0(1);                                        Leu, 7.6(8);           Lys, 3.9(4).    __________________________________________________________________________

Similarly, Compounds 22, 23 and 28 were prepared following thisprocedure.

    __________________________________________________________________________    Compound 22:    AVSEIQFLHN LGKHLSSLRR RELLEKLLEK LHNX-NH.sub.2 (SEQ ID NO:36)    (X = homoserine)    __________________________________________________________________________    Ala       Val          Ser             Glu                Ile                   Gln                      Phe                         Leu                            His                               Asn                                  Leu                                     Gly                                        Lys                                           His                                              Leu    1           5              10             15    Ser       Ser          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Asn             hSer NH.sub.2 (SEQ ID NO:36)    __________________________________________________________________________    Physical Data: m.p. 69.4-128° c.    α!.sub.D.sup.25   -43.93    (c 0.15, H.sub.2 O)    FAB (C.sub.179 H.sub.302 N.sub.56 O.sub.49):  M+H!.sup.+  4022.9    AAA:   Asx, 2.0(2);                    Glx, 4.9(5);                              Ser, 2.6(3);                                        His, 2.8(3);           Gly, 1.0(1);                    Ala, 1.0(1);                              Arg, 3.0(3);                                        Val, 1.0(1);           Phe, 1.0(1);                    Ile, 0.9(1);                              Leu, 8.8(9);                                        Lys, 3.4(3);    __________________________________________________________________________

    __________________________________________________________________________    Compound 23:    AVSEIQFLHN KGKHLSSLRR RELLEKLLEK LHNX-NH.sub.2 (SEQ ID NO:37)    (X = homoserine)    __________________________________________________________________________    Ala       Val          Ser             Glu                Ile                   Gln                      Phe                         Leu                            His                               Asn                                  Lys                                     Gly                                        Lys                                           His                                              Leu    1           5              10             15    Ser       Ser          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Asn             hSer NH.sub.2 (SEQ ID NO:37)    __________________________________________________________________________    Physical Data: m.p. 87.1-142.1° C.    α!.sub.D.sup.25    -52.14 (c 0.41, H.sub.2 O)    FAB (C.sub.179 H.sub.303 N.sub.57 O.sub.49):  M+H!.sup.+  4038    AAA:   Asx, 2.1(2);                    Glx, 4.9(5);                              Ser, 2.7(3);                                        His, 2.8(3);           Gly, 1.0(1);                    hSer, 1.0(1);                              Ala, 1.0(1);                                        Arg, 3.0(3);           Val, 1.1(1);                    Phe, 0.9(1);                              Ile, 0.9(1);                                        Leu, 7.9(8);           Lys, 3.7(4).    __________________________________________________________________________

    __________________________________________________________________________    Compound 28:    AVSEHQLLHD KGKSIQDLRR RELLRLLER LHTAGRRX-NH.sub.2 (SEQ ID NO:42)    (X = homoserine)    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                                           Glu                                              Arg                20             25             30    Leu       His          Thr             Ala                Gly                   Arg                      Arg                         hSer NH.sub.2 (SEQ ID NO:42)                35    __________________________________________________________________________    Physical Data: m.p. 80° C.    α!.sub.D.sup.25   -48.64 (c    0.09, H.sub.2 O)    FAB (C.sub.193 H.sub.334 N.sub.70 O.sub.56):  M+H!.sup.+  4530.0    AAA:   Asx, 2.2(2);                    Glx, 6.1(6);                              Ser, 1.7(2);                                        His, 3.0(3);           Gly, 1.9(2);                    hSer, 1.0(1);                              Thr, 1.0(1);                                        Ala, 2.1(2);           Arg, 7.2(7);                    Val, 0.8(1);                              Ile, 1.0(1);                                        Leu, 8.4(8);           Lys, 2.1(2).    __________________________________________________________________________

The homoserine alkylamides were similarly prepared from the homoserinelactone by dissolving it in DMF containing an excess of thecorresponding alkylamine. After stirring at room temperature for severaldays (the reaction was monitored by analytical HPLC) the mixture wasevaporated to dryness and the peptide purified by preparative HPLC.Representative homoserine alkylamides are Compounds 55 and 56.

    __________________________________________________________________________    Compound 55:    AVSEHOLLHD KGKSIQDLRR RELLEKLLEK LHTX-NHCH.sub.2 CH.sub.3 (SEQ ID NO:69)    (X = homoserine)    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             hSer NHCH.sub.2 CH.sub.3 (SEQ ID NO:69)    __________________________________________________________________________    Physical Data: m.p. (not determined)    α!.sub.D.sup.25   (not    determined)    FAB (C.sub.178 H.sub.306 N.sub.56 O.sub.52):  M+H!.sup.+  4063.0    AAA:   Asx, 2.1(2);                    Glx, 5.8(6);                              Ser, 1.7(2);                                        His, 3.1(3);           Gly, 0.9(1);                    Thr, 1.0(1);                              Ala, 0.9(1);                                        Arg, 3.0(3);           Val, 1.1(1);                    Ile, 1.0(1);                              Leu, 8.4(8);                                        Lys, 3.7(4);           hSer, 0.9(1).    __________________________________________________________________________

    __________________________________________________________________________    Compound 56:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTX-NHCH.sub.2 CH.sub.2 C.sub.6 H.sub.5    (SEQ ID NO:70)    (X = homoserine)    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             hSer NHCH.sub.2 CH.sub.2 C.sub.6 H.sub.5 (SEQ ID    __________________________________________________________________________             NO:70)    Physical Data: m.p. (not determined)    α!.sub.D.sup.25   (not    determined)    FAB (C.sub.184 H.sub.310 N.sub.56 O.sub.52):  M+H!.sup.+  4138.8    AAA:   Asx, 2.0(2);                    Glx, 5.9(6);                              Ser, 1.7(2);                                        His, 2.9(3);           Gly, 0.9(1);                    Thr, 1.0(1);                              Ala, 0.9(1);                                        Arg, 3.0(3);           Val, 1.0(1);                    Ile, 0.9(1);                              Leu, 8.0(8);                                        Lys, 4.1(4);           hSer, 0.9 (1).    __________________________________________________________________________

EXAMPLE IV

The cesium salt of N.sup.α -Boc-N.sup.γ -Fmoc-L-2,4-diaminobutyric acidwas attached to Merrifield resin (DMF, 50° C., 48 hrs) and used in asolid-phase synthesis in place of the Boc-Ala resin as in Example I.Upon completion of the synthesis the peptide was treated with 20%piperidine in DMF at room temperature for 30 minutes to remove the sidechain Fmoc protecting group. The protected peptide spontaneouslycyclized to the lactam thereby cleaving itself from the resin. Thesolution was filtered from the resin and evaporated under vacuum toleave on oil. The residue was treated with liquid HF as in Example I toyield the crude unprotected peptide. The peptide was treated andpurified as in Example I.

    __________________________________________________________________________    Compound 49:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTX (SEQ ID NO:63)    (X = L-2,4-diaminobutyryl lactam)    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             L-2,4-diaminobutyryl lactam (SEQ ID NO:63)    __________________________________________________________________________    Physical Data: m.p. 161-181° C.    α!.sub.D.sup.25   -48.38    (c 0.25, H.sub.2 O)    FAB (C.sub.176 H.sub.300 N.sub.56 O.sub.51):  M+H!.sup.+  4016.8    AAA:   Asx, 2.1(2);                    Glx, 6.3(6);                              Ser, 1.7(2);                                        His, 3.3(3);           Gly, 1.1(1);                    Thr, 1.0(1);                              Ala, 2.1(2);                                        Arg, 2.9(3);           Val, 0.9(1);                    Ile, 0.9(1);                              Leu, 8.0(8);                                        Lys, 3.8(4).    __________________________________________________________________________

EXAMPLE V

An aqueous solution of the homoserine lactone analog from Example II wastreated with porcine liver esterase (Sigma Chemical Company, St. Louis,Mo.). The hydrolysis of the lactone to the C-terminal homoserine wasmonitored by analytical HPLC. When the hydrolysis was judged to becomplete the material was purified by preparative HPLC as in Example I.

    __________________________________________________________________________    Compound 37:    AVSEHQLLHD KGKSIQDLRR RELLEKLLEK LHTX-OH (SEQ ID NO:51)    (X = homoserine)    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             hSer OH (SEQ ID NO:51)    __________________________________________________________________________    Physical Data: m.p. (not determined)    α!.sub.D.sup.25   (not    determined)    FAB (C.sub.176 H.sub.301 N.sub.55 O.sub.53):  M+H!.sup.+  4035.1    AAA:   Asx, 2.1(2);                    Glx, 5.9(6);                              Ser, 2.0(2);                                        His, 3.1(3);           Gly, 0.8(1);                    hSer, 0.8(1);                              Thr, 1.0(1);                                        Ala, 1.0(1);           Arg, 3.0(3);                    Val, 1.3(1);                              Ile, 1.0(1);                                        Leu, 8.1(8);           Lys, 3.8(4).    __________________________________________________________________________

EXAMPLE VI

Following Example I, the protected peptide-resinBocAVS(Bzl)E(OBz)H(Bom)QLLHD(OBzl)R(Ts)GR(Ts)S(Bzl)IQD(OBz)LR(Ts)R(Ts)E(OBz)LLE(OBzl)R(Ts)LLK(Fmoc)R(Ts)LH(Bom)T(Bzl)A-O-PAMwas synthesized on a 0.35 mmol scale. All N.sup.α groups were protectedwith t-butoxycarbonyl (Boc); side chain protecting groups were asindicated. After completion of the synthesis, the peptide resin wastreated with 50 mL of 20% piperidine in dimethylformamide (DMF) at roomtemperature for 30 minutes to remove the fluorenylmethoxycarbonyl (Fmoc)protecting group on lysine. The resin was washed successively with DMF,MeOH, CH₂ Cl₂ and dried to give 1.6 g partially protected peptide. 0.8 g(0.175 mmol) of the partially protected peptide was acylated on lysinewith 0.44 g (0.3 mmol) of methoxydi(ethyleneoxy) acetic acid PEG(2)CH₂COOH! in the presence of 0.16 g (0.3 mmol)benzotriazolyloxy-tris(pyrrolidino)phosphonium hexafluorophosphate(PyBop) and 0.067 g (0.525 mmol) of diisopropylethyl amine (DIEA) in 20mL DMF at room temperature for 5 hrs. After 5 hrs., the resin wasfiltered and washed successively with DMF, MeOH and CH₂ Cl₂. Theacylation step was repeated twice until negative ninhydrin result on theresin was obtained. The final peptide was cleaved from the resin withremoval of the side chain protecting groups and purification as inExample I; 100 mgs of Compound 19 were obtained.

    __________________________________________________________________________    Compound 19    AVSEHQLLHDRGRSIQDLRRRELLERLLKRLHTA-OH (SEQ ID NO:18)    CH.sub.3 O(CH.sub.2 CH.sub.2 O).sub.2 CH.sub.2 C═O    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Arg                                     Gly                                        Arg                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                            20          25    Lys(COCH.sub.2 PEG2)             Arg                Leu                   His                      Thr                         Ala                            OH (SEQ ID NO:18)                         30    __________________________________________________________________________    Physical Data: m.p. 145-195° C.    α!.sub.D.sup.25    -44.60° (c 0.2, H.sub.2 O)    FAB (C.sub.183 H.sub.316 N.sub.64 O.sub.54):  M+H!.sup.+  4276.2    AAA:   Asp, 2.1(2);                    Glu, 5.0(5);                              Ser, 1.6(2);                                        His, 2.9(3);           Gly, 0.9(1);                    Thr, 1.9(2);                              Arg, 7.1(7);                                        Val, 1.1(1);           Ile, 1.0(1);                    Leu, 8.0(8);                              Lys, 0.9(1).    __________________________________________________________________________

EXAMPLE VII

Peptide was synthesized, cleaved and purified in the same manner as inExample IV except 2-methoxypoly(ethyleneoxy) acetic acid PEG(5000)CH₂CO₂ H! was used as the acylating agent. 0.8 g (0.175 mmol) of partiallyprotected peptide yielded 300 mgs of pure Compound 20.

    __________________________________________________________________________    Compound 20    AVSEHQLLHDRGRSIQDLRRRELLERLLKRLHTA-OH (SEQ ID NO: 19)    CH.sub.3 O(CH.sub.2 CH.sub.2 O).sub.110 CH.sub.2 C═O    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Arg                                     Gly                                        Arg                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Arg                                     Leu                                        Leu                20             25    Lys(COCH.sub.2 PEG5000)                Arg                   Leu                      His                         Thr                            Ala                               OH (SEQ ID NO:19).                30    __________________________________________________________________________    Physical Data: m.p. 105° C.    α!.sub.D.sup.25    -22.95° (c 0.11, 50% aq. HOAc)    AAA:   Asp, 2.0(2);                    Glu, 4.8(5);                              Ser, 1.6(2);                                        His, 2.6(3);           Gly, 1.1(1);                    Thr, 1.1(1);                              Arg, 7.3(7);                                        Val, 0.8(1);           Ile, 0.9(1);                    Leu, 8.3(8);                              Lys, 1.1(1);                                        Ala, 1.8(2).    __________________________________________________________________________

EXAMPLE VIII Synthesis of hPTHrp(1-34) analog gene

A synthetic gene coding for the hPTHrp(1-34) analog Compound 4 (SEQ IDNO:9) was designed having the nucleotide sequence and enzyme restrictionsites shown in FIG. 1. The requisite oligodeoxynucleotides were preparedwith a DNA synthesizer (Milligen/Biosearch) using the phosphoramiditeprocess of Sinha, et al., Nucleic Acid Research 12, 4539-4557 (1984).After deprotection, the crude oligonucleotides were purified by gelelectrophoresis on preparative 150 polyacrylamide gels. Theoligonucleotides were located with UV, excised from the gel, desaltedover Waters c18 Sep-pak® cartridges, and lyophilized.

Amplification via polymerase chain reaction (PCR) was carried out on aPerkin-Elmer Cetus thermal cycler, with 25 cycles of: 94° C. for 1minute, 50° C. for 2 minutes, and 72° C. for 3 minutes, using reagents,including Taq polymerase, from the "GeneAmp" DNA amplification kit(Perkin-Elmer Cetus).

Two overlapping oligonucleotides, an 88mer (2 μg), PTH3,

    __________________________________________________________________________    (SEQ ID NO:31)    CCTCTAGATC            TCCGCGGCGC                    TAGC                        ATG GCT                               GTT                                  TCT                                     GAA                                        CAT                                           CAG                                              45                                  Met                                     Ala                                        Val                                           Ser                                              Glu    His       Gln                            1           5    CTG       CTT          CAT             GAC                AAA                   GGT                      AAA                         TCG                            ATT                               CAA                                  GAT                                     CTG                                        AGA                                           CGT                                              C 88    Leu       Leu          His             Asp                Lys                   Gly                      Lys                         Ser                            Ile                               Gln                                  Asp                                     Leu                                        Arg                                           Arg             10             15             20    and an anti-sense 90mer (2 μg), PTH4 (SEQ ID NO:32):    CCTCGAAGCT            TATGCATCAT                    TATCTAGA                            CAT                               AGT                                  ATG                                     CAG                                        CTT                                           TTC                                              46                      Met                         Thr                            His                               Leu                                  Lys                                     Glu                               30    AAG       CAG          TTT             CTC                CAG                   CAG                      CTC                         GCG                            ACG                               TCT                                  CAG                                     ATC                                        TTG                                           AAT                                              88    Leu       Leu          Lys             Glu                Leu                   Leu                      Glu                         Arg                            Arg                               Arg                                  Leu                                     Asp                                        Gln                                           Ile          25          20          15    CG 90,    __________________________________________________________________________

were prepared as the template DNA sequence for the hPTHrp (1-34) analoggene. Utilizing the two flanking primers, PTHPCR1: CCTCTAGATC TCCGCGGCGCTAG (SEQ ID NO:33) and PTHPCR2: CCTCGAAGCT TATGCATCAT TATC (SEQ IDNO:34), the entire gene was amplified by PCR. The amplified DNA productswere purified by gel electrophoresis on 40 NuSieve® agarose gel. Theband containing the synthetic hPTHrp(1-34) analog gene, approximately150 bases in length, was excised from the gel and approximately 200 ngof DNA isolated by Elu-Quik® glass gel DNA extraction (Schleicher &Schuell, Keene, N.H.).

EXAMPLE IX Molecular Cloning of an hPTHrp(1-34)1 Analog Gene

To subclone the hPTHrp(1-34) analog gene of Example VI, 200 ng of theamplified DNA was isolated and cut by restriction enzymes HinD III andSac II. As shown in FIG. 2, the DNA was ligated to 2 μg TrpLE 18 Prot(Ile³,Pro⁵) plasmid previously cleaved with Hind III and Sac II.

The resulting plasmid, TrpLE 18 hPTHrp(1-34)1, containing one copy ofthe hPTHrp(1-34) analog gene was then transformed into competent E. coliHB 101 cells (CLONTECH, Palo Alto, Calif.). Transformants were subjectedto PCR analysis to verify insertion. Transformed cell colonies wereselected and boiled in 200 μL of water for 5 minutes; 2 μL weresubjected to PCR with two primers flanking the insert. The PCR productwas then analyzed on 1% agarose gel to confirm the presence of one copyof the hPTHrp(1-34) gene insert. TrpLE 18 hPTHrp(1-34)1 construct wasthen verified by DNA sequencing on an automated DNA sequencer (AppliedBiosystems Model 373A, Foster City, Calif.) using the vendor's Dye DeoxyTerminator Sequencing kit.

EXAMPLE X Construction of a Trp LE 18 vector containing multiple copiesof the hPTHrp(1-34) analog gene

Unique Nhe I and Xba I restriction sites are located near the beginningand end of the hPTHrp(1-34) analog gene sequence. These two sites, whichrecognize different sequences, but produce identical single strandcohesive termini, allow the construction of multiple copy hPTHrp (1-34)genes within the Trp LE 18 vector.

The strategy for constructing repeated hPTHrp(1-34) sequences in tandemis outlined in FIG. 3. In separate reactions, 5 μg of plasmid Trp LE 18hPTHrp(1-34)1 containing a single copy of the gene was cleaved with BamHI+Nhe I and Xba I+Bam HI. From each digest, about 300 ng of thefragment containing the hPTHrp(1-34) analog gene was isolated. These twofragments were mixed and ligated to form the Trp LE 18 hPTHrp(1-34)2plasmid. This plasmid was used to transform competent E. coli HB 101cells. Sizing of the transformed PCR products on 1% agarose gel was usedto determine the presence of two copies of the hPTHrp(1-34) gene insert.TrpLE 18 hPTHrp(1-34)2 containing two copies of the gene was thenconfirmed by DNA sequencing. The correct fusion of the two hPTHrp(1-34)genes results in the elimination of Nhe I and Xba I sites at thejunction. This makes the remaining Xba I and Nhe I sites flanking thetandem genes unique.

By repeating this process the final plasmid Trp LE 18 hPTHrp(1-34)4containing four copies of the hPTHrp(1-34) gene was constructed, asshown in FIG. 4. The sequence of Trp LE 18 hPTHrp(1-34)4 was found to becorrect by DNA sequence analysis.

EXAMPLE XI Expression and Purification of Trp LE 18 hPTHrp(1-34)4Induction of the Trp LE 18 hPTHrp(1-34)4

A starter culture of 50 mL of LB culture media, J. H. Miller,"Experiments in Molecular Genetics," p.431 (1972), incorporated hereinby reference, containing 5 μg/mL ampicillin and 10 μg/mL tryptophan, wasinoculated with E. coli cells containing Trp LE 18 hPTHrp(1-34)4plasmid, and grown overnight at 37° C. with vigorous shaking to an A₅₅₀of about 6. Two liters of LB culture media for production werepre-warmed to 37° C. and seeded with 20 mL of the starter culture togive an A₅₅₀ of about 0.06. The culture was then grown with vigorousshaking to an A₅₅₀ of between 0.6 and 0.8, whereupon 2 mL of a 10 mg/mLsolution of indole acrylic acid (IAA) was added. Growth was continuedwith good aeration for about 16 hr to a final A₅₅₀ of about 6(typically, between 4 and 10). The cells were concentrated bycentrifugation and resuspended in 500 mL of 50 mM Tris-HCl, pH 7.5, 0.1mM EDTA buffer solution (Tris buffer).

The suspension was sonicated using a Heat Systems-Ultrasonics, Inc.model 220F sonicator (equipped with a 3/4" horn) operated at 50% of fullcapacity to avoid overheating.

To determine the extent of induction, the whole cells were analyzed bySDS-PAGE. The gene products derived from the TrpLE 18 hPTHrp(1-34)4construct were seen as a major band of the predicted MW of approximately17,000. This accounts for as much as 100 of the total cellular protein.

Isolation of the Fusion Protein

The cell lysate was centrifuged for 15 min. at about 3600×g to pelletthe Trp LE 18 hPTHrp(1-34)4 fusion protein; the supernatant wasdiscarded. The pellet was resuspended in 200 mL Tris buffer. (typically40-80 A₅₅₀ /mL).

EXAMPLE XII Processing of the Fusion Protein and Purification ofhomo-Serlactone hPTHro(1-34) peptide

Cleavage of the methionine residues flanking the hPTHrp(1-34) multimericfusion protein with CNBr releases the desired homo-SerlactonehPTHrp(1-34) polypeptide, which was purified as described below.

CNBr Treatment of Fusion Protein

The washed pellet of TrpLE 18 hPTHrp(1-34)4 fusion protein wasresuspended by gently stirring in 60 mL of 700 formic acid (about 20mg/mL total protein; typically material from 1000 A₅₅₀ units of cells isdissolved in 3 mL). A few drops of octanol were added and N₂ bubbledthrough the solution for 20 minutes before adding 5.5 g CNBr. Thisreaction was allowed to proceed for 6 hours at 25° C. before an equalvolume of 50:50 MeOH:H₂ O was mixed with the sample and subsequentlyremoved by rotary evaporation. After 2 to 4 repetitions of this process,the bulk of the formic acid and CNBr were essentially removed. Thesample was then evaporated to dryness, redissolved in 200 mL water andlyophilized for storage.

Purification of homo-Serlactone hPTHrp(1-34)

The CNBr cleaved supernatant was dialyzed against 50 mM KH₂ PO₄ pH 6.5for 24 hours with multiple changes. During dialysis, pH was maintainedat 6.5. After dialysis, the precipitates were removed by high speedcentrifugation. The supernatant was clarified through a Gelman 0.45μfilter device (Acrodisc 4184).

Cation Exchange Chromatography

Initial purification was accomplished by cation exchange chromatographyon a Bio-Gel TSK-SP-5PW HPLC column (21.5×150 mm). Chromatographicconditions for a flow rate of 8 mL/min and a yield of approximately 12mg of highly purified homo-Serlactone hPTHrp(1-34) peptide were:

1. Column equilibration in 50 mM KH₂ PO₄, pH 6.5

2. Load 10 mL clarified supernatant (approximately 1.5 L culture brothor 2.4 g inclusion).

3. Wash column with 50 mM KH₂ PO₄ pH 6.5 containing 50 mM NaCl untilbaseline is stabilized.

4. Elute column with 50 mM KH₂ PO₄ pH 6.5 containing 90 mM NaCl. Collectfractions for about 45 minutes.

5. Analyze the 90 mM NaCl fractions for homo-Serlactone hPTHrp(1-34)content by C18 HPLC before pooling and storage.

Reverse Phase HPLC Chromatography

A reverse phase Poros R/H 4.6×100 mm column (Perseptive Biosystems,Cambridge, Mass.) was used for the final purification step. Thechromatographic conditions were as follows:

    ______________________________________    Mobile phase A: 0.1% trifluoroacetic acid (TFA)/water    B: 0.1% trifluoroacetic acid (TFA)/CH.sub.3 CN    TIME            FLOW     % B    ______________________________________      0 min         4 ml/min 15    5.0 min         4 ml/min 40    5.2 min         4 ml/min 100    6.8 min         4 ml/min 100    7.0 min         4 ml/min 15    ______________________________________

Retention time of the homo-Serlactone hPTHrp(1-34), Compound 4, wasapproximately 2.943 minutes. The purified peptide was approximately 98%pure as determined by mass spectroscopy.

EXAMPLE XIII

The compounds of this invention were evaluated for their effect on bonemass in ovariectomized rats, generally in accord with the procedures ofGunness-Hey and Hock, Metab. Bone Dis. 5:177 181 (1984).

Adult Sprague-Dawley female rats were acclimatized, weight grouped (n=9,10 or 12), and subjected to bilateral ovariectomy (OVX) or sham surgery.Dosing was initiated 17 days after surgery and continued for 20 days.Test compound was administered subcutaneously once a day in 2% ratserum/saline vehicle.

After 20 days of dosing, the rats were sacrificed and the right femursexcised. The femurs were cut in half and the distal half femurs (DHF)were further separated into trabecular bone (TB) and cortical bone (CB)by drilling out the trabeculae. Calcium was extracted and measured byCalcette calcium analyzer and expressed as mean bone Ca in mg/DHF/100 gbody weight.

The two sample t-test was used to compare OVX and sham groups. One-wayANOVA was used to compare OVX groups, followed by Fisher's LSD multiplecomparison to compare each treatment group to vehicle.

Ovariectomy induced substantial total bone loss, primarily fromtrabecular bone. Total bone calcium was 47 to 54% lower than forsham-operated controls.

bPTH(1-34) and hPTHrp(1-34) at 80 μg/kg/day provided statisticallysignificant increases in total bone calcium for treated OVX rats,ranging from 53 to 95% and 18 to 40%, respectively; however, there wasno significant increase in cortical bone calcium.

Compounds of this invention, dosed at 80 μg/kg/day, increased total bonecalcium by from 66 to 138% and trabecular calcium by from 87 to 128%.Cortical bone calcium, trabecular thickness, and bone volume were alsosignificantly increased over untreated OVX controls.

In this assay, the following compounds were tested:

    ______________________________________                           Trabecular Bone                                       Total Bone                           Calcium     Calcium    Compound       n (#    (% increase (% increase    Number         of tests)                           over OVX)   over OVX)    ______________________________________    Compound 1 (SEQ ID NO:7)                   6       101-128%    88-138%    Compound 2 (SEQ ID NO:6)                   3        87-102%    66-114%    Compound 4 (SEQ ID NO:9)                   3       --          88-114%    ______________________________________

In similar studies, ovariectomized rats were dosed for 5, 10 and 20days, at 40 ug/kg/day, with the following results:

    ______________________________________                                       Total Bone                                       Calcium    Compound        n (#    # of Days (d)                                       (% increase    Number          of tests)                            of dosing  over OVX)    ______________________________________    Compound 1 (SEQ ID NO:7)                    3       20 d       73-109%    Compound 4 (SEQ ID NO:9)                    5       20 d       79-105%    Compound 4 (SEQ ID NO:9)                    1       10 d       79%    Compound 49 (SEQ ID NO:63)                    1       10 d       93%    Compound 4 (SEQ ID NO:9)                    1        5 d       55%    Compound 42 (SEQ ID NO:56)                    1        5 d       60%    ______________________________________

EXAMPLE XIV TOXICITY

In the above Example XI, no toxic effects were observed with thecompounds of the invention.

    __________________________________________________________________________    SEQUENCE LISTING    (1) GENERAL INFORMATION:    (iii) NUMBER OF SEQUENCES: 86    (2) INFORMATION FOR SEQ ID NO:1:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:1:    SerValSerGluIleGlnLeuMetHisAsnLeuGlyLysHisLeuAsn    151015    SerMetGluArgValGluTrpLeuArgLysLysLeuGlnAspValHis    202530    AsnPhe    (2) INFORMATION FOR SEQ ID NO:2:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:2:    AlaValSerGluIleGlnPheMetHisAsnLeuGlyLysHisLeuSer    151015    SerMetGluArgValGluTrpLeuArgLysLysLeuGlnAspValHis    202530    AsnPhe    (2) INFORMATION FOR SEQ ID NO:3:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:3:    SerValSerGluIleGlnLeuMetHisAsnLeuGlyLysHisLeuSer    151015    SerLeuGluArgValGluTrpLeuArgLysLysLeuGlnAspValHis    202530    AsnPhe    (2) INFORMATION FOR SEQ ID NO:4:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:4:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgPhePheLeuHisHisLeuIleAlaGluIleHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:5:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:5:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuArgLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:6:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:6:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:7:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:7:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:8:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 34    (D) OTHER INFORMATION: /note= "Xaa34 = homoserine"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:8:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrXaa    (2) INFORMATION FOR SEQ ID NO:9:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 34    (D) OTHER INFORMATION: /note= "Xaa34 = homoserine lactone"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:9:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrXaa    (2) INFORMATION FOR SEQ ID NO:10:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 37 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:10:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAlaGlyArgArg    35    (2) INFORMATION FOR SEQ ID NO:11:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:11:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuLys    202530    GluLeu    (2) INFORMATION FOR SEQ ID NO:12:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:12:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuAlaArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:13:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:13:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgAlaGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:14:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:14:    AlaValSerGluAlaGlnLeuLeuHisAspLeuGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    AlaLeu    (2) INFORMATION FOR SEQ ID NO:15:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:15:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluArgLeuLeuGluArgLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:16:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:16:    AlaValSerGluHisGlnLeuLeuHisAspArgGlyArgSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluArgLeuLeuGluArgLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:17:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:17:    AlaValSerGluHisGlnLeuLeuHisAspArgGlyArgSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluArgLeuLeuLysArgLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:18:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 29    (D) OTHER INFORMATION: /note= "Xaa29 =    lysine- (OCCH2(OCH2CH2)2OCH3)"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:18:    AlaValSerGluHisGlnLeuLeuHisAspArgGlyArgSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluArgLeuLeuXaaArgLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:19:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 29    (D) OTHER INFORMATION: /note= "Xaa29 =    lysine- (OCCH2(OCH2CH2)110OCH3"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:19:    AlaValSerGluHisGlnLeuLeuHisAspArgGlyArgSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluArgLeuLeuXaaArgLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:20:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:20:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgAlaLeuAlaGluAlaLeuAlaGluAlaLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:21:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:21:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgSerLeuLeuSerSerLeuLeuSerSerLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:22:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:22:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgAlaPheTyrAspLysValAlaGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO:23:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 8    (D) OTHER INFORMATION: /note= "Xaa8 = norleucine"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 18    (D) OTHER INFORMATION: /note= "Xaa18 = norleucine"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:23:    AlaValSerGluIleGlnPheXaaHisAsnLeuGlyLysHisLeuSer    151015    SerXaaGluArgValGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    AsnTyr    (2) INFORMATION FOR SEQ ID NO:24:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 8    (D) OTHER INFORMATION: /note= "Xaa8 = norleucine"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 18    (D) OTHER INFORMATION: /note= "Xaa18 = norleucine"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:24:    AlaValSerGluIleGlnPheXaaHisAsnLeuGlyLysHisLeuSer    151015    SerXaaArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    AsnTyr    (2) INFORMATION FOR SEQ ID NO:25:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 35 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:25:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrMetAla    35    (2) INFORMATION FOR SEQ ID NO:26:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 10 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: internal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 8    (D) OTHER INFORMATION: /note= "Xaa8 = glutamic acid or    arginine"    (ix) FEATURE:    (A) NAME/KEY: Region    (B) LOCATION: 1..10    (D) OTHER INFORMATION: /note= "Sequence 26 is embedded at    positions 22 to 31 of sequences 5, 6, 7, 8, 9, 10,    11, 12, 13, and 14."    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:26:    GluLeuLeuGluLysLeuLeuXaaLysLeu    1510    (2) INFORMATION FOR SEQ ID NO:27:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 10 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: internal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 8    (D) OTHER INFORMATION: /note= "Xaa8 = glutamic acid,    lysine, or lysine-(OCCH2PEGX)"    (ix) FEATURE:    (A) NAME/KEY: Region    (B) LOCATION: 1..10    (D) OTHER INFORMATION: /note= "Sequence 27 is embedded at    positions 22 to 31 of sequences 15, 16, 17, 18,    and 19. "    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:27:    GluLeuLeuGluArgLeuLeuXaaArgLeu    1510    (2) INFORMATION FOR SEQ ID NO:28:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 10 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: internal    (ix) FEATURE:    (A) NAME/KEY: Peptide    (B) LOCATION: 1..10    (D) OTHER INFORMATION: /note= "Sequence 28 is embedded at    positions 22 to 31 of sequence 20."    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:28:    AlaLeuAlaGluAlaLeuAlaGluAlaLeu    1510    (2) INFORMATION FOR SEQ ID NO:29:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 10 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: internal    (ix) FEATURE:    (A) NAME/KEY: Peptide    (B) LOCATION: 1..10    (D) OTHER INFORMATION: /note= "Sequence 29 is embedded at    positions 22 to 31 of sequence 21."    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:29:    SerLeuLeuSerSerLeuLeuSerSerLeu    1510    (2) INFORMATION FOR SEQ ID NO:30:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 10 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: internal    (ix) FEATURE:    (A) NAME/KEY: Peptide    (B) LOCATION: 1..10    (D) OTHER INFORMATION: /note= "Sequence 30 is embedded at    positions 22 to 31 of sequence 22."    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:30:    AlaPheTyrAspLysValAlaGluLysLeu    1510    (2) INFORMATION FOR SEQ ID NO:31:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 88 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: cDNA    (iii) HYPOTHETICAL: NO    (iv) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:31:    CCTCTAGATCTCCGCGGCGCTAGCATGGCTGTTTCTGAACATCAGCTGCTTCATGACAAA60    GGTAAATCGATTCAAGATCTGAGACGTC88    (2) INFORMATION FOR SEQ ID NO:32:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 90 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: cDNA    (iii) HYPOTHETICAL: NO    (iv) ANTI-SENSE: YES    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:32:    CCTCGAAGCTTATGCATCATTATCTAGACATAGTATGCAGCTTTTCAAGCAGTTTCTCCA60    GCAGCTCGCGACGTCTCAGATCTTGAATCG90    (2) INFORMATION FOR SEQ ID NO:33:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 23 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: cDNA    (iii) HYPOTHETICAL: NO    (iv) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:33:    CCTCTAGATCTCCGCGCGCTAGC23    (2) INFORMATION FOR SEQ ID NO:34:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 24 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: cDNA    (iii) HYPOTHETICAL: NO    (iv) ANTI-SENSE: YES    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:34:    CCTCGAAGCTTATGCATCATTATC24    (2) INFORMATION FOR SEQ ID NO: 35:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 35:    AlaValSerGluIleGlnPheLeuHisAsnLeuGlyLysHisLeuSer    151015    SerLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    AsnTyr    (2) INFORMATION FOR SEQ ID NO: 36:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 34    (D) OTHER INFORMATION: /note= "Xaa is homoserine"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 36:    AlaValSerGluIleGlnPheLeuHisAsnLeuGlyLysHisLeuSer    151015    SerLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    AsnXaa    (2) INFORMATION FOR SEQ ID NO: 37:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 34    (D) OTHER INFORMATION: /note= "Xaa is homoserine"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 37:    AlaValSerGluIleGlnPheLeuHisAsnLysGlyLysHisLeuSer    151015    SerLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    AsnXaa    (2) INFORMATION FOR SEQ ID NO: 38:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 38:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuLysLeuLysGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO: 39:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 39:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluArgLeuLeuGluArgLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO: 40:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 35 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 40:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluArgLeuLeuGluArgLeuHis    202530    ThrAlaPro    35    (2) INFORMATION FOR SEQ ID NO: 41:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 37 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 41:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluArgLeuLeuGluArgLeuHis    202530    ThrAlaGlyArgArg    35    (2) INFORMATION FOR SEQ ID NO: 42:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 38 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 38    (D) OTHER INFORMATION: /note= "Xaa is homoserine"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 42:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluArgLeuLeuGluArgLeuHis    202530    ThrAlaGlyArgArgXaa    35    (2) INFORMATION FOR SEQ ID NO: 43:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 43:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrTyr    (2) INFORMATION FOR SEQ ID NO: 44:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 44:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyTyrSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO: 45:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 45:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyCysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO: 46:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 13    (D) OTHER INFORMATION: /note= "Xaa is    Cys(CH- 2CONH(CH-2)-2NH(biotinyl))"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 46:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyXaaSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO: 47:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 13    (D) OTHER INFORMATION: /note= "Xaa is    Lys(7- dimethylamino-2-oxo-2H-1-benxopyran-4-acety    l)"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 47:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyXaaSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO: 48:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 35 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 48:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAlaGly    35    (2) INFORMATION FOR SEQ ID NO: 49:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 33 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 4    (D) OTHER INFORMATION: /note= "Xaa4 is Glu(OCH-3)"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 10    (D) OTHER INFORMATION: /note= "Xaa10 is Asp(OCH-3)"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 17    (D) OTHER INFORMATION: /note= "Xaa17 is Asp(OCH-3)"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 22    (D) OTHER INFORMATION: /note= "Xaa22 is Glu(OCH3)"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 25    (D) OTHER INFORMATION: /note= "Xaa25 is Glu(OCH-3)"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 29    (D) OTHER INFORMATION: /note= "Xaa29 is Glu(OCH-3)"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 49:    AlaValSerXaaHisGlnLeuLeuHisXaaLysGlyLysSerIleGln    151015    XaaLeuArgArgArgXaaLeuLeuXaaLysLeuLeuXaaLysLeuHis    202530    Ala    (2) INFORMATION FOR SEQ ID NO: 50:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 33 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 4    (D) OTHER INFORMATION: /note= "Xaa4 is Glu(OCH-3)"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 10    (D) OTHER INFORMATION: /note= "Xaa10 is Asp(OCH-3)"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 17    (D) OTHER INFORMATION: /note= "Xaa17 is Asp(OCH-3)"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 22    (D) OTHER INFORMATION: /note= "Xaa22 is Glu(OCH-3)"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 25    (D) OTHER INFORMATION: /note= "Xaa25 is Glu(OCH-3)"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 29    (D) OTHER INFORMATION: /note= "Xaa29 is Glu(OCH-3)"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 50:    AlaValSerXaaHisGlnLeuLeuHisXaaLysGlyLysSerIleGln    151015    XaaLeuArgArgArgXaaLeuLeuXaaLysLeuLeuXaaLysLeuHis    202530    Ala    (2) INFORMATION FOR SEQ ID NO: 51:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 34    (D) OTHER INFORMATION: /note= "Xaa is homoserine"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 51:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrXaa    (2) INFORMATION FOR SEQ ID NO: 52:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 35 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 52:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAlaPro    35    (2) INFORMATION FOR SEQ ID NO: 53:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 53:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrPro    (2) INFORMATION FOR SEQ ID NO: 54:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 54:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrPro    (2) INFORMATION FOR SEQ ID NO: 55:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 33 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 55:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    Pro    (2) INFORMATION FOR SEQ ID NO: 56:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 32 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 56:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuPro    202530    (2) INFORMATION FOR SEQ ID NO: 57:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 37 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 57:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrArgSerAlaTrp    35    (2) INFORMATION FOR SEQ ID NO: 58:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 42 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 58:    AlaValSerGluHisGlnLeuLeuHisAspArgGlyArgSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluArgLeuLeuGluArgLeuHis    202530    ThrAlaGlyArgArgThrArgSerAlaTrp    3540    (2) INFORMATION FOR SEQ ID NO: 59:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 42 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 59:    AlaValSerGluHisGlnLeuLeuHisAspArgGlyArgSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluArgLeuLeuGluArgLeuHis    202530    ThrArgGlyArgArgThrArgSerAlaTrp    3540    (2) INFORMATION FOR SEQ ID NO: 60:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 42 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 13    (D) OTHER INFORMATION: /note= "Xaa13 is    Lys(dihydrocinnamoyl)"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 60:    AlaValSerGluHisGlnLeuLeuHisAspArgGlyXaaSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluArgLeuLeuGluArgLeuHis    202530    ThrArgGlyArgArgThrArgSerAlaTrp    3540    (2) INFORMATION FOR SEQ ID NO: 61:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 8    (D) OTHER INFORMATION: /note= "Leu8 is Norleucine"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 18    (D) OTHER INFORMATION: /note= "Leu18 is Norleucine"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 61:    AlaValSerGluIleGlnPheLeuHisAsnLeuGlyLysHisLeuSer    151015    SerLeuThrArgSerAlaTrpLeuArgLysLysLeuGlnAspValHis    202530    AsnTyr    (2) INFORMATION FOR SEQ ID NO: 62:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 35 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 62:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrMetAla    35    (2) INFORMATION FOR SEQ ID NO: 63:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 33 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 33    (D) OTHER INFORMATION: /note= "Xaa is Thr    1,4- diaminobutyryl lactam"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 63:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    Xaa    (2) INFORMATION FOR SEQ ID NO: 64:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 64:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgPhePheLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO: 65:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 65:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuHisLysLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO: 66:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 66:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluHisLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO: 67:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 67:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuIleAlaLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO: 68:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 68:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluGluIleHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO: 69:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 34    (D) OTHER INFORMATION: /note= "Xaa is homoserine"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 69:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrXaa    (2) INFORMATION FOR SEQ ID NO: 70:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 34    (D) OTHER INFORMATION: /note= "Xaa is homoserine"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 70:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrXaa    (2) INFORMATION FOR SEQ ID NO: 71:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: both    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 71:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAla    (2) INFORMATION FOR SEQ ID NO: 72:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 37 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 72:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrArgSerAlaTrp    35    (2) INFORMATION FOR SEQ ID NO: 73:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 36 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 36    (D) OTHER INFORMATION: /note= "Xaa is Ala    3-(2- naphthyl)-L-alanine"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 73:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrArgSerXaa    35    (2) INFORMATION FOR SEQ ID NO: 74:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 37 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 74:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAlaSerAlaTrp    35    (2) INFORMATION FOR SEQ ID NO: 75:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 38 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 75:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAlaGluIleArgAla    35    (2) INFORMATION FOR SEQ ID NO: 76:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 37 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 76:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAlaGluIleArg    35    (2) INFORMATION FOR SEQ ID NO: 77:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 36 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 77:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAlaGluIle    35    (2) INFORMATION FOR SEQ ID NO: 78:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 35 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 78:    AlaValSerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGln    151015    AspLeuArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHis    202530    ThrAlaGlu    35    (2) INFORMATION FOR SEQ ID NO: 79:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 34 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: N-terminal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 8    (D) OTHER INFORMATION: /note= "Leu8 is Norleucine"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 18    (D) OTHER INFORMATION: /note= "Leu18 is Norleucine"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 34    (D) OTHER INFORMATION: /note= "Xaa is homoserine lactone"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 79:    AlaValSerGluIleGlnPheLeuHisAsnLysGlyLysHisLeuSer    151015    SerLeuGluArgValGluTrpLeuArgLysLysLeuGlnAspValHis    202530    AsnXaa    (2) INFORMATION FOR SEQ ID NO: 80:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 32 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: internal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 80:    SerGluHisGlnLeuLeuHisAspLysGlyLysSerIleGlnAspLeu    151015    ArgArgArgGluLeuLeuGluLysLeuLeuGluLysLeuHisThrAla    202530    (2) INFORMATION FOR SEQ ID NO: 81:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 28 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: internal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 81:    LeuLeuHisAspLysGlyLysSerIleGlnAspLeuArgArgArgGlu    151015    LeuLeuGluLysLeuLeuGluLysLeuHisThrAla    2025    (2) INFORMATION FOR SEQ ID NO: 82:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 27 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: internal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 82:    LeuHisAspLysGlyLysSerIleGlnAspLeuArgArgArgGluLeu    151015    LeuGluLysLeuLeuGluLysLeuHisThrAla    2025    (2) INFORMATION FOR SEQ ID NO: 83:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 41 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: internal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 83:    SerGluHisGlnLeuLeuHisAspArgGlyArgSerIleGlnAspLeu    151015    ArgArgArgGluLeuLeuGluArgLeuLeuGluArgLeuHisLeuHis    202530    ArgGlyArgArgThrArgSerAlaTrp    3540    (2) INFORMATION FOR SEQ ID NO: 84:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 35 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: internal    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 84:    LeuLeuHisAspArgGlyArgSerIleGlnAspLeuArgArgArgGlu    151015    LeuLeuGluArgLeuLeuGluArgLeuHisAlaGlyArgArgThrArg    202530    SerAlaTrp    35    (2) INFORMATION FOR SEQ ID NO:85:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 10 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: internal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 1 and 4    (D) OTHER INFORMATION: /note= "Xaa1 and Xaa4 = Glu,    Glu(OCH3), His, or Phe"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 2    (D) OTHER INFORMATION: /note= "Xaa2 = Leu or Phe"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 5    (D) OTHER INFORMATION: /note= "Xaa5 = Lys or His"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 7 and 10    (D) OTHER INFORMATION: /note= "Xaa7 and Xaa10 =    Leu or Ile"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 8    (D) OTHER INFORMATION: /note= "Xaa8 = Ala, Arg, or Glu"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 9    (D) OTHER INFORMATION: /note= "Xaa9 = Lys or Glu"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:85:    XaaXaaLeuXaaXaaLeuXaaXaaXaaXaa    1510    (2) INFORMATION FOR SEQ ID NO:86:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 10 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (iii) HYPOTHETICAL: NO    (v) FRAGMENT TYPE: internal    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 1 and 4    (D) OTHER INFORMATION: /note= "Xaa1 and Xaa4 = Glu,    Glu(OCH3), His, or Phe"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 2    (D) OTHER INFORMATION: /note= "Xaa2 = Leu or Phe"    (ix) FEATURE:    (A) NAME/KEY: Modified-site    (B) LOCATION: 8    (D) OTHER INFORMATION: /note= "Xaa8 = Glu, Lys or    Lys(COCH2PEGX)"    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:86:    XaaXaaLeuXaaArgLeuLeuXaaArgLeu    1510    __________________________________________________________________________

We claim:
 1. A modified parathyroid hormone (PTH) or parathyroid hormonerelated peptide (PTHrP) having bone mss restoring activity which differsfrom naturally occurring PTH or PTHrP by changes comprisingsubstitutions at one or more of positions Xaa(22-31>, wherein the Xaa(22-31)substitutions are selected from the group consisting of SEQ IDNos. (26, 27, 28, 29, 30, 85 and 86).
 2. A polypeptide of claim 1wherein Xaa²²⁻³¹ is (SEQ ID NO:26).
 3. The polypeptide of claim 2 whichis:

    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       Pro          NH.sub.2             (SEQ ID NO:56).    __________________________________________________________________________


4. The polypeptide of claim 2 which is:

    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             1,4-diaminobutyryl lactam (SEQ ID NO:63).    __________________________________________________________________________


5. The polypeptide of claim 2 which is:

    __________________________________________________________________________    Ala       Val          Ser             Glu                His                   Gln                      Leu                         Leu                            His                               Asp                                  Lys                                     Gly                                        Lys                                           Ser                                              Ile    1           5              10             15    Gln       Asp          Leu             Arg                Arg                   Arg                      Glu                         Leu                            Leu                               Glu                                  Lys                                     Leu                                        Leu                                           Glu                                              Lys                20             25             30    Leu       His          Thr             hSerlac (SEQ ID NO:9).    __________________________________________________________________________


6. A polypeptide of claim 2 wherein Xaa¹¹ and Xaa¹³ are both Lys; andone of Xaa¹⁹ and Xaa²¹ is Arg and the other is Ala.
 7. A polypeptide ofclaim 2 wherein one of Xaa¹¹ and Xaa¹³ is Leu and the other is Lys; andXaa¹⁹ and Xaa²¹ are both Arg.
 8. A polypeptide of claim 1 whereinXaa²²⁻³¹ is (SEQ ID NO:27).
 9. A polypeptide of claim 8 wherein Xaa¹¹and Xaa¹³ are both Lys or both Arg; and Xaa¹⁹ and Xaa²¹ are both Arg.10. A polypeptide of claim 1 wherein Xaa²²⁻³¹ is (SEQ ID NO:28).
 11. Apolypeptide of claim 1 wherein Xaa²²⁻³¹ is (SEQ ID NO:29).
 12. Apolypeptide of claim 1 wherein Xaa²²⁻³¹ is (SEQ ID NO:30).
 13. Apharmaceutical composition for the prevention or treatment of conditionscharacterized by decreases in bone mass comprising an effective bonemass increasing amount of a polypeptide of claim 1, or salt thereof, anda pharmaceutically acceptable carrier.
 14. A pharmaceutical compositionin unit dosage form, for treating conditions characterized by decreasesin bone mass, comprising from about 0.1 μg to about 50 μg of apolypeptide of claim 1, or salt thereof, and a pharmaceuticallyacceptable carrier.